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The Pap smear has engendered a tremendous amount of media coverage in recent years.  In spite of the emotional and regulatory issues surrounding the test, two facts remain.  First, the Pap smear is a screening test.   It identifies cells which may indicate a precancerous or cancerous process.   However, any abnormal smear must be followed with a careful clinical examination and biopsy.  Second, the Pap smear is one of the great success stories in medicine.  In fifty years, it has largely helped to reduce the incidence of cervical cancer from one of the leading causes of death in women in the United States to the eighth most common cancer (about 4500 deaths annually).  It is estimated that there has been a 75% reduction in cervical cancer deaths since the introduction of the Pap smear. Over 50 million cervical cytologic Pap smears are performed in the United States each year.

An average slide contains 50,000-300,000 cells which the cytotechnologist, who is screening the slide, must review. To make this task even more difficult, there are numerous pre-analytical variables which can make this daunting task even more difficult, such as air-drying, low cellularity, excessive blood and bacteria. It is a tribute to the skill and diagnostic accuracy of these cytotechnologists that the Pap smear has evolved into such an effective screen for cervical cancer.


How Accurate is a Pap Smear?  
How is a Pap Smear Obtained?  
What is an adequate specimen?  
Bethesda Classification  

Comparative Studies
Inform HPV
Thin Prep
Autopap (Surepath)

Costs of the Pap Smear  
Cytologic Variants  
Differential Diagnosis

Endometrial cells
Hyperchromatic crowded groups
Glandular cells in post-hysterectomy patients
Perimenopausal Cells (PM Cells)
Small blue cells of Tamoxifen
High grade Vulvar Intraepithelial Neoplasia (VIN III)

Prognosis and Treatment  
Commonly Used Terms  
Internet Links  


How Accurate is a Pap Smear?

There is no test in medicine which is 100% accurate. Even if all the proper collection techniques, specimen handling, and screening procedures are utilized, there will still be a false negative (missed lesion) rate of at least 4%. Up to 2/3 of false negative Pap smear result from factors related to the collection procedure. However, the natural history of cervical dysplasias and carcinomas is such that there is a long time interval (years) from dysplasia to invasive carcinoma. This is why a yearly screening is so important because even if this occurs, the chance of a lesion being missed is very low.

Double Slide Viewing as a Cytology Quality Improvement Initiative

Stephen S. Raab, MD, etal.

Am J Clin Pathol 2006;125:526-533
Abstract quote

Few studies have measured the effect of pre–sign out double viewing of cytology cases as a means to decrease error. Three Agency for Healthcare Research and Quality–funded project sites performed pre–sign out double viewing of 431 pulmonary cytology cases.

Two-step or more differences in diagnosis were arbitrated as interpretive errors, and the effect of double viewing was measured by comparing the frequency of cytologic-histologic correlation–detected errors in the previous 2 years with the double-viewing period. The number of interpretive errors detected by double viewing for the 3 institutions was 2.7%, 0%, and 1.9%, respectively. Double viewing did not lower the frequency of cytologic-histologic correlation false-negative errors.

We conclude that double viewing detects errors in up to 1 of every 37 cases and that biases in the double-viewing process limit error detection.

Quality indices in a cervicovaginal cytology service: before and after laboratory accreditation.

Tan KB, Chang SA, Soh VC, Thamboo TP, Nilsson B, Chan NH.

Department of Pathology, National University of Singapore, National University Hospital, Singapore.
Arch Pathol Lab Med. 2004 Mar;128(3):303-7. Abstract quote  

CONTEXT: Quality assurance practices contribute to the effectiveness of cervical screening and are formalized by participation in a laboratory accreditation program.

OBJECTIVE: To identify changes in the quality indices of our cervicovaginal cytology service preceding and following laboratory accreditation by the College of American Pathologists in 2000.

DESIGN: Cervicovaginal cytology quality indices for 2001 (postaccreditation) were compared with those of 1997 (preaccreditation). Performances in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology (PAP) for the years 1999-2002 were analyzed.

RESULTS: A comparison between data for 1997 and 2001 shows the following: (a) a higher inadequacy rate (1.3% vs 0.7%; P <.001) in 2001; (b) maintenance of a low atypical squamous cells of undetermined significance-squamous intraepithelial lesion ratio (0.79 and 0.76, respectively); (c) overall positive predictive values of positive cytology of 82% and 87%, respectively; (d) relatively few changes to the original cytologic diagnoses following review of significant cytohistologic discrepancies (4 cases and 2 cases, respectively); and (e) a higher subsequent positive yield of squamous intraepithelial lesions following atypical squamous cells of undetermined significance diagnoses in 2001 (41% vs 19%; P =.02). The performance of the laboratory and cytotechnologists in the PAP program showed maintenance of a high standard with almost no major discrepancies recorded.

CONCLUSIONS: An increased awareness of quality-related issues and participation in intradepartmental consultation/diagnostic seminars, all part of the accreditation process, have very likely contributed to the modest improvements identified in the cytology service. Future challenges include increases in workload with the anticipated launch of Singapore's national cervical screening program and adaptation to the emerging cervical screening technologies.

A practical problem with calculating the false-negative rate of Papanicolaou smear interpretation by rescreening negative cases alone.

Renshaw AA.

Department of Pathology, Baptist Hospital of Miami, Miami, Florida 33176, USA.

Cancer 1999 Dec 25;87(6):351-3 Abstract quote

BACKGROUND: Rescreening negative Papanicolaou (Pap) smears alone is the most commonly employed method of determining the false-negative rate (FNR) or the false-negative proportion for a laboratory. Acceptable FNRs have been proposed, and the number of slides needed to be rescreened to demonstrate a statistically significant difference in FNRs has been determined. The authors sought to determine the range of FNRs this method can measure and, by implication, the value of this method.

METHODS: A literature review and an analysis of the FNRs this method can generate was performed.

RESULTS: If one assumes that the FNR of review is the same as that of initial screening, the maximum measured FNR is only 25%, even with a true FNR of anywhere from 0-100%. In fact, as a laboratory's FNR increases over 50%, the measured FNR decreases back to zero. This range of FNRs corresponds very closely to the published range of FNRs of 1.6-28%. Because many authorities believe that 5% may be the lowest achievable FNR, the entire possible range of measured FNRs is only 5-25%. In this setting, a statistically significant difference of 20% is meaningless, and a statistically significant difference of 10% can only be achieved by laboratories with an initial FNR of less than 15%, and actual changes in FNR that are much greater than 10%.

CONCLUSIONS: FNRs determined by review of negative smears without abnormal smears generate unreliable and potentially seriously misleading results. Current methodologies exist for more accurately determining the FNR of Pap smear screening by incorporating abnormal smears into the review process. There is little justification for further review of negative Pap smears alone as a method for determining the FNR of a laboratory.

Estimating the percentage of Papanicolaou smears that can be reproducibly identified: modeling Papanicolaou smear interpretation based on multiple blinded rescreenings.

Renshaw AA.

Department of Pathology, Baptist Hospital of Miami, Miami, Florida 33176, USA.

Cancer 2001 Aug 25;93(4):241-5 Abstract quote

BACKGROUND: Multiple blinded rescreenings of Papanicolaou (Pap) smears for litigation purposes is based on the assumption that a subset of Pap smears can be reproducibly identified. The size of this subset is not known.

METHODS: To estimate the size of the subset of Pap smears that can be reproducibly identified, a model was constructed based on the results of repeated blinded screenings in the AutoPap Primary Screening System Trial. Additional analysis came from data in the literature.

RESULTS: Routine and AutoPap-assisted screening both have a detection rate for all detected abnormal cases of < 50%. Models with only two subsets or types of slides each with a different detection rate correlated well with the available data. Data from multiple rapid reviews strongly supported the existence of additional definable subsets. Although the percentage of cases with an expected detection rate of 100% in a three-subset model might have been as high as 30% of the abnormal cases detected in a single review, all estimates that included a second subset of slides with at least a 50% detection rate limited the percentage of slides in the 100% sensitive subset of slides to < 2% of all abnormal slides and < 6% of all abnormal slides detected by a single screening.

CONCLUSIONS: Repeated screenings of Pap smears allowed more accurate models of the sensitivity of Pap-smear screening and the overall incidence of abnormal cases. The data strongly supported the existence of multiple subsets of Pap smears, which can be defined by repeated blinded rescreenings. The percentage of slides that can be reproducibly identified was small.

Papanicolaou tests diagnosed as atypical by a cytotechnologist and downgraded to benign by a pathologist: a measure of laboratory quality.

Condel JL, Mahood LK, Grzybicki DM, Sturgis CD, Raab SS.

Department of Pathology and Laboratory Medicine, Allegheny General Hospital, Pittsburgh, PA 15212, USA.

Am J Clin Pathol 2002 Apr;117(4):534-40 Abstract quote

Follow-up of Papanicolaou (Pap) tests diagnosed as atypical squamous cells of undetermined significance (ASCUS) or atypical glandular cells of undetermined significance (AGUS) by a cytotechnologist and downgraded to benign by a pathologist has not been measured.

Squamous intraepithelial lesion (SIL) follow-up rates were obtained for Pap tests diagnosed as ASCUS (288) or AGUS (94) and downgraded to benign and for Pap tests diagnosed as repair (231). Statistically significant associations were seen between 7 cytotechnologists and between 7 pathologists and ASCUS, AGUS, downgraded ASCUS, and downgraded AGUS rates. The percentage of downgraded ASCUS cases compared with all ASCUS cases per pathologist ranged from 4.8% to 43.7%. Statistically significant associations between pathologists and SIL follow-up rates for downgraded ASCUS diagnoses were seen. The SIL follow-up rate for repair (7.9%) was similar to that for a downgraded ASCUS (11.0%) or AGUS (7.3%).

The parameters of downgraded ASCU and AGUS Pap test interpretations are good quality indicators of individual performance and overall laboratory quality.

Cytohistologic discrepancies: a means to improve pathology practice and patient outcomes.

Clary KM, Silverman JF, Liu Y, Sturgis CD, Grzybicki DM, Mahood LK, Raab SS.

Department of Pathology and Laboratory Medicine, Allegheny General Hospital, Pittsburgh, PA 15212, USA.

Am J Clin Pathol 2002 Apr;117(4):567-73 Abstract quote

The use of cytohistologic discrepancies to investigate and reduce error seldom is studied.

All gynecologic discrepancies (n = 283; 0.87% and 7.37% of all cytologic and histologic cases, respectively) and nongynecologic discrepancies (n = 146; 2.26% and 0.44% of all cytologic and histologic cases, respectively) for 26 months were classified as sampling or interpretive. Specimen type and pathologist discrepancy percentages, effect of discrepancies on patient outcome, and interobserver agreement of discrepancies were evaluated. Discrepancies were interpretive in 67% and 34% of gynecologic and nongynecologic cases, respectively. Statistically significant associations were seen between individual pathologist and discrepancy percentages. Breast (1.2%) and bronchial (0.8%) cytologic diagnoses had the highest discrepancy percentages. The kappa scores ranged from 0.02 to 0.45 for pairwise agreement of discrepant cases.

Of nongynecologic interpretive discrepancies available for review, 63% (27/43) and 14% (6/43) were of no or minor clinical significance, respectively. Cytohistologic correlation is a useful tool to monitor performance and to identify specimen types prone to error.

How Is A Pap Smear Obtained?

The collection procedure actually begins with appropriate instruction of the patient regarding the test. A Pap test should be obtained: Annually after the age of 18 or after the beginning of sexual activity During the second half of the menstrual cycle-i.e., at least two weeks after the start of one menstrual period and before the start of the next menstrual period Without intercourse during the 24 hours prior to the test Without douching during the 24 hours prior to the test

The health care provider is responsible for providing accurate information including:
Patient name
Patient age
Last menstrual period
Pregnancy history
History of hormone use
History of IUD use
Risk factors
Previous abnormal Pap smears
Relevant clinical information-.e.g., abnormal bleeding, discharge, pelvic pain

The specific collection procedure utilized will depend on the type of information required or the specific indication for performing the Pap smear. The goal of the actual collection procedure is to produce an adequate, evaluable smear of cellular material from the vagina and/or cervix which can be submitted to the laboratory, along with appropriate clinical information, to be stained and evaluated in accordance with the indication for the test. In

Single-slide Pap smear: an acceptable alternative to the double-slide Pap smear.

Quackenbush SR.

Pathology Interpretation, P.C., Pennsylvania, Ottumwa, IA 52501, USA.

Diagn Cytopathol 1999 May;20(5):317-20 Abstract quote

Pap smears obtained with Ayre spatula and cytobrush by four experienced private practice gynecologists were submitted to the cytology laboratory on either two slides or one slide. For each physician, 500 consecutive cases were studied for each of 4 consecutive years. Compared to the first 2 years (double-slide cases), the rates of abnormal smears increased from 4.28 to 5.58% while the number of unsatisfactory smears decreased from 1.83 to 0.98% for the years with all single-slide cases. These changes were not statistically significant. Follow-up biopsy findings were not different between the groups. Similar findings were seen when the medical community that submits smears to this laboratory began to use primarily a single-slide technique.

The daily screening workload in the laboratory decreased 29% as mostly one-slide smears were submitted despite an increase of 10% in the number of cases. The double-slide method for Pap smears can be safely replaced by a single-slide method.

What Is An Adequate Specimen?

Adequate numbers of squamous epithelial cells present Evidence that the transformation zone was sampled (i.e., the presence of endocervical cells on the smear) Spread in a relatively even monolayer Epithelial cells not obscured by blood, inflammatory cells, or foreign material such as lubricant or talc appropriately preserved.

Clinical Significance of Unsatisfactory Conventional Pap Smears Owing to Inadequate Squamous Cellularity Defined by the Bethesda 2001 Criterion

Amy L. Adams, MD, etal.
Am J Clin Pathol 2005;123:738-743 Abstract quote

To determine the incidence of clinically significant lesions in long-term follow-up after a diagnosis of inadequate squamous cellularity using former and new criteria, we reviewed conventional Papanicolaou (Pap) smears (January-December 1998) for adequacy based on the Bethesda System 2001 criterion. Of 23,302 Pap smears evaluated in our laboratory, 114 (0.489%) were classified as unsatisfactory and 245 (1.051%) as "satisfactory but limited by" based on the 10% rule. Follow-up information for 5 years was obtained for 172 patients without a concurrent cervical epithelial abnormality: 25 (14.5%) had squamous abnormalities (atypical squamous cells, 22; low-grade squamous intraepithelial lesion, 2; and high-grade squamous intraepithelial lesion, 1).

With the Bethesda System 2001 criterion, 167 (97.1%) of 172 smears had inadequate squamous cellularity and 5 (2.9%) were adequate. No differences in the incidence of squamous abnormalities detected on follow-up were noted between patients with unsatisfactory Pap smears owing to inadequate squamous cellularity and patients with satisfactory and negative smears.

Our findings raise the question whether patients with unsatisfactory Pap smears and a negative history of gynecologic diseases require repeated Pap smears within 2 to 4 months as suggested by the American Society for Colposcopy and Cervical Pathology guideline.
Interobserver Variability in Assessing Adequacy of the Squamous Component in Conventional Cervicovaginal Smears

Matthew V. Sheffield, MD,1 Aylin Simsir, MD,2 Lynya Talley, PhD,3 A. Janie Roberson, SCT(ASCP),1 Paul A. Elgert, CT(ASCP)CMIAC,2, and David C. Chhieng, MD

Am J Clin Pathol 2003;119:367-373 Abstract quote

We compared the interobserver reproducibility of estimating the adequacy of the squamous component of conventional Papanicolaou (Pap) smears using traditional and newly proposed criteria. Forty conventional Pap smears with varying degrees of squamous cellularity were reviewed by 13 observers who evaluated adequacy (satisfactory vs unsatisfactory) based on the traditional criterion of estimating 10% slide coverage.

After being introduced to the new criterion and the reference images, the observers reevaluated adequacy on the same set of smears, using the new criterion and the reference images. With the original criterion of 10% slide coverage, 15 smears had a unanimous designation; the overall kappa value was 0.49 (P < .001). With the newly proposed adequacy criterion and reference images, 17 smears had a unanimous designation; the overall kappa value was 0.60 (P < .001). The difference in the kappa correlation coefficients was statistically significant (P = .007).

While traditional and newly proposed criteria resulted in fair interobserver agreement, it seemed that the newly proposed criterion, along with the use of reference images, for evaluating adequacy of the squamous component of conventional Pap smears results in better interobserver reproducibility.

Dual Sampling of the Endocervix and Its Impact on AutoCyte Prep Endocervical Adequacy

Sarah J. Day, CT(ASCP)
Eric L. Deszo
and Gregory G. Freund, MD

Am J Clin Pathol 2002;118:41-46 Abstract quote

We compared satisfactory for evaluation but limited by (limited by) and unsatisfactory gynecologic cytologic diagnoses for samples collected by conventional smearing with those generated with the AutoCyte Prep in a population with a historic squamous intraepithelial lesion (SIL) rate of less than 1%.

Results from 18,819 AutoCyte Preps were compared with 53,835 conventional cervical smears. Furthermore, 23 women ages 18 to 65 years undergoing annual Papanicolaou tests underwent sequential sampling with the AutoCyte Prep and the Surgipath C-E brush. Comparison of the AutoCyte Prep with conventional cytologic diagnoses revealed the following: unsatisfactory rate, down 97%; limited by rate, down 67%; low-grade SIL rate, up 86%; cervical cancer rate, up 300%; and high-grade SIL rate, unchanged. Examination of unsatisfactory and limited by cases for the AutoCyte Prep showed that 88% were due to absence of endocervical cells (ECs). Dual sampling showed no improvement in EC recovery over the AutoCyte collection device. Compared with conventional Papanicolaou smears, the AutoCyte Prep significantly decreased the rate of unsatisfactory and limited by specimens while increasing low-grade SIL and cancer detection and EC recovery.

The majority of limited by specimens with the AutoCyte Prep were due to absence of ECs, but use of a brush-type device for better endocervical sampling did not enhance EC recovery.

ASCCP Patient Management Guidelines*
Pap Test Specimen Adequacy and Quality Indicators

Diane D. Davey, MD, Chair,
R. Marshall Austin, MD, PhD,
George Birdsong, MD
Henry W. Buck, MD,4 J. Thomas Cox, MD
Teresa M. Darragh, MD
Paul A. Elgert, CT(ASCP)
Vivien Hanson, MD
Michael R. Henry, MD
and Jeffrey Waldman, MD

Am J Clin Pathol 2002;118:714-718 Abstract quote

Our objective was to provide management guidelines according to Papanicolaou (Pap) test specimen adequacy based on literature review and expert opinion. A task force named by the American Society for Colposcopy and Cervical Pathology (ASCCP) conducted a literature review and discussed appropriate management. The Steering Committee of the ASCCP and other experts reviewed the guidelines.

The guidelines recommend a repeated Pap test in 12 months for most women undergoing routine annual/biennial screening if the current Pap test is negative but either lacks an endocervical/ transformation zone component or is partially obscured. Indications for considering an earlier repeat are also provided.

The preferred management for unsatisfactory Pap tests is a repeated Pap test within a short interval of 2 to 4 months. The management guidelines will help promote optimal and uniform follow-up of women according to Pap test specimen adequacy.

Endocervical Status Is Not Predictive of the Incidence of Cervical Cancer in the Years After Negative Smears

Anita B. Bos, MSc,1 Marjolein van Ballegooijen, PhD,1 M. Elske van den Akker-van Marle, MSc,1 Antonius G.J.M. Hanselaar, PhD,2 Gerrit J. van Oortmarssen, PhD,1 and J. Dik F. Habbema, PhD

Am J Clin Pathol 2001;115:851-855 Abstract quote

The clinical relevance of the lack of endocervical cells was never well established in a longitudinal study with histologically proven cervical cancer as an end point.

From the Dutch Network and National Database for Pathology, results for all negative smears obtained in 1990 and 1991 in the Netherlands were retrieved, as were data for all cytologic and histologic examinations performed after the negative smears before April 1998. There were no significant differences between the proportion of preinvasive lesions (cervical intraepithelial neoplasia 1, 2, and 3) detected after negative smears without endocervical cells compared with negative smears with endocervical cells. The proportion of women in whom invasive cancer developed was the same in both groups.

These data suggest there is no reason to advise women with negative smears without endocervical cells to undergo an additional smear.

What Terms Does a Pathologist Use to Diagnose Abnormal Cells on a Pap Smear? (The Bethesda Classification)

For many years, diagnoses of cytological abnormalities was based upon a class system. For example, Class II Pap smears were benign reactive changes. In 1988, a multinational workshop was convened and formulation a new classification system known as the Bethesda Classification. This introduced terms such as ASCUS (Atypical squamous cells of undetermined significance) and LSIL (Low grade squamous intraepithelial lesion). In 1991, this classification was revised. In 2001, a third workshop was held which further refined the classification system. This system acknowledges the introduction of new diagnostic techniques such as thin layer (liquid based) cytology as well as ancillary testing such as DNA probes for HPV. It eliminates the previous category of benign reactive changes, placing it in the category of negative for intraepithelial lesion or malignancy. It also eliminates the term ASCUS and replaces it with ASC (Atypical squamous cells). It further refines these changes as either of undetermined significance or cannot exclude high grade squamous intraepithelial cells.


NEGATIVE FOR INTRAEPITHELIAL LESION OR MALIGNACY   When there is no cellular evidence of neoplasia, state this in the General Categorization above and/or in the Interpretation /Result section of the report, whether or not there are organisms or other non-neoplastic findings
ORGANISMS Trichomonas vaginalis
Fungal organisms morphologically consistent with Candida spp.
Shift in flora suggestive of bacterial vaginosis
Bacteria morphologically consistent with Actinomyces spp.
Cellular changes consistent with Herpes simplex virus
OTHER ENDOMETRIAL CELLS In a woman >/= 40 years
Specify if negative for squamous intraepithelial lesion
Atypical squamous cells of undetermined significance (ASC-US)  
Atypical squamous cells-cannot exclude HSIL (ASC-H)  
Low grade squamous intraepithelial lesion (LSIL) Encompassing HPV/mild dysplasia/CIN 1
High grade squamous intraepithelial lesion (HSIL) Encompassing moderate and severe dysplasia, CIS/CIN2 and CIN 3

May add with features suspicious for invasion (if invasion is suspected)
Squamous cell carcinoma  
Atypical endocervical cells NOS or specify in comments
Atypical endometrial cells NOS or specify in comments
Atypical glandular cells NOS or specify in comments
Atypical endocervical cells, favor neoplastic  
Atypical glandular cells, favor neoplastic  
Endocervical adenocarcinoma in situ  
Adenocarcinoma Endocervical
Not otherwise specified (NOS)
Bethesda 2001 implementation and reporting rates: 2003 practices of participants in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology.

Davey DD, Neal MH, Wilbur DC, Colgan TJ, Styer PE, Mody DR.

Department of Pathology and Laboratory Medicine, University of Kentucky, Lexington 40526-0298, USA.
Arch Pathol Lab Med. 2004 Nov;128(11):1224-9. Abstract quote  

CONTEXT: The 2001 Bethesda System for the reporting of cervical cytology specimens and the use of new liquid-based and human papillomavirus testing technologies have led to changes in cervical cytology reporting practices.

OBJECTIVES: To analyze current laboratory reporting practices using Bethesda 2001 terminology and to compare results with previous survey data from 1996.

DESIGN: Questionnaire survey mailed to 1751 laboratories in mid-2003.

PARTICIPANTS: Laboratories enrolled in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology.

RESULTS: Of the 759 responding laboratories, most (85.5%) had implemented Bethesda 2001 terminology, and the majority had adopted major changes, such as elimination of the benign cellular changes category and the satisfactory but limited category. The median reporting rate for low-grade squamous intraepithelial lesion was 2.1%, compared to a 1996 median rate of 1.6%, but the increase was confined to liquid-based preparations. Reporting rates for high-grade squamous intraepithelial lesion (median, 0.5%) and atypical squamous cells (ASC) had changed little. Most ASC cases were subclassified as "undetermined significance" (median, 3.9%) with ASC, cannot exclude high-grade SIL accounting for a minority (median, 0.2%). The median ratio of ASC to squamous intraepithelial lesions and carcinomas (SIL+) was 1.4 and was lower than the 1996 median ratio of 2.0. Median reporting rates for squamous abnormalities for 2002 were noted to be significantly higher for liquid-based preparations than for conventional smears, while median ASC/SIL+ ratios were lower. Most laboratories offer human papillomavirus testing, but almost half (47%) of laboratories do not know the percentage of positive testing results in patients with ASC. CONCLUSIONS: Most laboratories have implemented Bethesda 2001 terminology. New criteria and liquid-based methods have led to an increase in low-grade squamous intraepithelial lesion reporting rates and a decrease in ASC/ SIL+ ratios when compared with 1996 data. Liquid-based preparations have higher median squamous intraepithelial lesion rates and lower ASC/SIL+ ratios than conventional smears.

What New Techniques Are Available to Assist the Pathologist?

Currently, there are a number of sophisticated tests which have been used as an adjunct to the Pap smear. These adjuvant tests may play a role in triaging Pap smears which are characterized as ASCUS (atypical squamous cells of undetermined significance) and LSIL (low grade squamous intraepithelial lesions). The ASCUS/LSIL Triage Study (ALTS) is a multicenter study comparing different strategies for management. Over 5000 women are enrolled with random assignment to one group receiving colposcopy with repeat Pap test every 6 months, one group with repeat Pap tests every 6 months, and one group receiving a Pap test plus an HPV test. Early reports have indicated that HPV is highly prevalent in women with LSIL, limiting the usefulness of the HPV testing in this population. Others have advocated that HPV negative ASCUS cases should be classified as benign.

One adjuvant test is the second generation Hybrid Capture II (HCII) from Digene (Gaithersburg, MD) utilizes a DNA binding assay. The specimens are obtained from a cervical swab and the material is subjected to a base which disrupts the virus and releases the target DNA. A specific RNA probe is added which binds to the target DNA forming a DNA:RNA hybrid. These hybrids are captured on a solid phase coated with antibodies specific for the DNA:RNA hybrids. These captured hybrids are then detected with antibodies conjugated to alkaline phosphatase which amplifies the signal 3000 fold. The bound alkaline phosphatase is detected by a chemiluminescent dioxetane substrate. When cleaved with alkaline phosphatase, the substrate produces light which is measured. This test is very accurate detecting 89% of high-grade lesions and 100% of cervical cancers in 8,554 women in a high risk population. This test is more sensitive than Pap smears (88.4% vs. 77.7%) but less specific (89.0% vs. 94.2%).

The Cytyc Corporation ThinPrep Pap Test creates a monolayer of cells allowing for greater visualization of cellular detail. The cell imaging device of Tripath's AutoPap uses a neural net computer program that screens the microscopic slide cells using complex algorithms and displaying the abnormal cells for final review by a cytotechnologist. These abnormal cells are then reviewed by a pathologist and a final diagnosis is rendered. This device also combines a monolayer technology to produce a comprehensive package.


Comparison of performance of conventional and ThinPrep gynecologic preparations in the College of American Pathologists Gynecologic Cytology Program.

Renshaw AA, Young NA, Birdsong GG, Styer PE, Davey DD, Mody DR, Colgan TJ.

College of American Pathologists Gynecologic Cytology Program, Northfield, Ill., USA.

Arch Pathol Lab Med. 2004 Jan;128(1):17-22. Abstract quote  

CONTEXT: Results of clinical trials suggest that interpretation of liquid-based cytology preparations is more accurate and is associated with less screening error than interpretation of conventional preparations.

OBJECTIVE: In this study, the performance of participants in interpreting ThinPrep (TP) preparations was compared with participants' performance on conventional Papanicolaou tests in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology (PAP).

DESIGN: The results of the PAP from the year 2002 were reviewed, and the discordancies to series and exact-match error rates for the 2 cytologic methods were compared.

RESULTS: For this study, a total of 89 815 interpretations from conventional smears and 20 886 interpretations from TP samples were analyzed. Overall, interpretations of TP preparations had both significantly fewer false-positive (1.6%) and false-negative (1.3%) rates than those of conventional smears (P =.001 and P =.02, respectively) for validated or validated-equivalent slides, as assessed by concordance with the correct diagnostic series. In this assessment of concordance to series, interpretations of educational TP and conventional preparations were similar, except for high-grade squamous intraepithelial lesion, in which the performance was significantly worse for educational TP preparations (false-negative rate of 8.1% vs 4.1% for conventional smears, P <.001). When interpretations were matched to the exact diagnosis, validated-equivalent TP preparations were generally more accurate for diagnoses in the 100 series and 200 series than were conventional smears. Notably, for the reference diagnosis of squamous cell carcinoma, the exact-match error rate on validated equivalent TP slides was significantly greater than that of conventional slides (44.5% vs 23.1%, P <.001). Interpretations of educational TP preparations also had a significantly higher error rate in matching to the exact reference diagnosis for squamous cell carcinoma (33.7% vs 22.8%, P =.007).

CONCLUSIONS: Overall, TP preparations in this program were associated with significantly lower error rates than conventional smears for both validated and educational cases. However, unlike the negative for intraepithelial lesion and malignancy, not otherwise specified, low-grade squamous intraepithelial lesion, and adenocarcinoma cytodiagnostic challenges, participants' responses indicated some difficulty in recognizing high-grade squamous intraepithelial lesion and squamous cell carcinoma.

Comparative analysis of conventional papanicolaou tests and a fluid-based thin-layer method.

Limaye A, Connor AJ, Huang X, Luff R.

Department of Anatomic Pathology, Quest Diagnostics Incorporated, Teterboro, NJ.


Arch Pathol Lab Med 2003 Feb;127(2):200-4 Abstract quote

Context.-A fluid-based, direct-to-vial method of thin-layer gynecologic cytology (ThinPrep Pap Test) is reported to be more effective than the conventional Papanicolaou test in the detection of squamous intraepithelial lesions.

Objective.-This retrospective analysis evaluated the validity of the findings on the thin-layer method using case material at a large independent laboratory and represented a comparison of performance of both methods over an identical period.

Methods.-Data for conventional and ThinPrep tests were compared for 2 periods. Period 1 included 1 421 080 conventional and 56 835 ThinPrep specimens, and period 2 included 564 270 conventional and 109 784 ThinPrep specimens. Squamous intraepithelial lesions were used to determine detection of disease. These 2 sets of data were also analyzed to eliminate effects of any selection bias toward ThinPrep for high-risk patients. Results.-Use of ThinPrep showed a greater than 100% increase in the detection rate of squamous intraepithelial lesions (1.3%-3.4% in period 1 and 1.3%-2.9% in period 2), which was statistically significant after correcting for selection bias. We also found a significant decrease in the false-negative proportion (57% in period 1 and 35% in period 2). There was a marked improvement (233%) in the detection of high-grade squamous intraepithelial lesions in high-risk cases and a decrease in the atypical squamous cells of undetermined significance to squamous intraepithelial lesion ratio from 3.1 to 1.5 in period 2.

Conclusion.-ThinPrep is better than the conventional Papanicolaou test in detecting squamous intraepithelial lesions and is a superior screening test in detection of precancerous changes of the cervix.

Cost-effectiveness of 3 methods to enhance the sensitivity of Papanicolaou testing.

Brown AD, Garber AM.

Department of Public Health and Primary Care, University of Oxford, England.

JAMA 1999 Jan 27;281(4):347-53 Abstracgt quote

CONTEXT: ThinPrep, AutoPap, and Papnet are 3 new technologies that increase the sensitivity and cost of cervical cancer screening.

OBJECTIVE: To estimate the cost-effectiveness of these technological enhancements to Papanicolaou (Pap) tests.

DESIGN: We estimated the increase in sensitivity from using these technologies by combining results of 8 studies meeting defined criteria. We used published literature and additional sources for cost estimates. To estimate overall cost-effectiveness, we applied a 9-state time-varying transition state model to these data and information about specific populations.

SETTING: A hypothetical program serving a cohort of 20- to 65-year-old women who begin screening at the same age and are representative of the US population.

RESULTS: The new technologies increased life expectancy by 5 hours to 1.6 days, varying with the technology and the frequency of screening. All 3 technologies also increased the cost per woman screened by $30 to $257 (1996 US dollars). AutoPap dominated ThinPrep in the base case. At each screening interval, AutoPap increased survival at the lowest cost. The cost per year of life saved rose from $7777 with quadrennial screening to $166000 with annual screening. Papnet produced more life-years at a higher cost per year of life saved. However, when used with triennial screening, each of them produced more life-years at lower cost than conventional Pap testing every 2 years. The cost-effectiveness ratio of each technology improved with increases in the prevalence of disease, decreases in the sensitivity of conventional Pap testing, and increases in the improvement in sensitivity produced by the technology.

CONCLUSIONS: Technologies to increase the sensitivity of Pap testing are more cost-effective when incorporated into infrequent screening. Increases in sensitivity and decreases in cost may eventually make each technology more cost-effective.

Comparison of ThinPrep and TriPath PREP liquid-based preparations in nongynecologic specimens: a pilot study.

Michael CW, McConnel J, Pecott J, Afify AM, Al-Khafaji B.

Department of Pathology, University of Michigan, Ann Arbor, Michigan 48109-0054, USA.

Diagn Cytopathol 2001 Sep;25(3):177-84 Abstract quote

ThinPrep (TP) and TriPath PREP (TriP) are two liquid-based cytologic preparations that produce a thin layer of cells.

This study compares the diagnostic accuracy and different cytomorphologic alterations produced by these preparations in nongynecologic specimens.

Samples from 10 urines (3 urothelial carcinomas and 7 negative), 4 positive serous fluids, and 7 fine-needle aspirates (FNAs) were prepared by both techniques. FNAs represented one each of: Hashimoto's thyroiditis (HT), hyperplastic colloid nodule (HCN), Hodgkin's lymphoma, liposarcoma, chondrosarcoma, squamous-cell carcinoma (SCC) metastatic to the lymph node, and carcinoid tumor. All 5 participants, none of whom had prior knowledge of the clinical history or histologic diagnosis, reviewed and interpreted the slides. Both techniques produced a clean background and were equally accurate in urines, serous fluids, and three FNAs. TriP was slightly more accurate in four FNAs: HCN and HT where colloid and lymphocytes were better represented, SCC where keratin and malignant cells were more readily identified among lymphocytes, and carcinoid which was easier to evaluate on TriP due to less cellular shrinkage and more dispersion of cells between aggregates. TP preparations had more cell shrinkage, and the chromatin was harder to evaluate. Both techniques produced artificial aggregations of lymphocytes, but TriP had a more evenly dispersed single-cell population between aggregates, rendering them easier to evaluate for atypia. TP produced fragmentation of large sheets that were flattened, while TriP contained larger branching sheets in a three-dimensional (3-D) configuration. TP produced a true monolayer of cells that were all spread at the same plane, while in TriP the cells were spread at slightly different planes, requiring frequent focusing of the viewed plane.

While both techniques are acceptable for diagnostic purposes, they both introduce new cytomorphologic alterations that pathologists need to recognize. TriP seems superior to TP in FNAs specimens where preservation of architecture and cellular integrity are important considerations.

Clinical and cost implications of new technologies for cervical cancer screening: the impact of test sensitivity.

Hutchinson ML, Berger BM, Farber FL.

Department of Pathology and Laboratory Medicine, Woimen and Infants' Hospital of Rhode Island, Providence, USA.

Am J Manag Care 2000 Jul;6(7):766-80 Abstract quote

OBJECTIVE: To compare the available techniques for cervical cancer screening, including several new technologies, using actual program utilization patterns.

STUDY DESIGN: Longitudinal cohort model.

PATIENTS AND METHODS: The model followed a cohort of 100,000 women who underwent screening from age 20 through 65 years. The model was run with a weighted average of screening intervals to model the actual utilization of the cervical cancer screening program in the United States.

RESULTS: The model demonstrated that new technologies with significantly increased test sensitivity have the potential to reduce the number of cancers by 45% to 60% depending on the screening frequency in fully compliant populations. At screening intervals of 2 years or more, these new technologies had cost-effectiveness ratios below $50,000 per life-year saved. Assuming existing utilization patterns, the model predicted there would be 13.2 cancers per year in the 100,000 women screened with the conventional Pap smear, and new technologies with increased test sensitivity could reduce the annual incidence to 9.5 cancers per 100,000 women screened.

CONCLUSIONS: The model suggests that to achieve further dramatic reduction in cervical cancer mortality, significant improvements in test sensitivity, as reflected in the new screening technologies, may be the most realistic and cost-effective approach.

Setting the target for a better cervical screening test: characteristics of a cost-effective test for cervical neoplasia screening.

Myers ER, McCrory DC, Subramanian S, McCall N, Nanda K, Datta S, Matchar DB.

Department of Obstetrics and Gynecology, Center for Clinical Health Policy Research-Evidence-Based Practice Center, Durham, North Carolina, USA.

Obstet Gynecol 2000 Nov;96(5 Pt 1):645-52 Abstract quote

OBJECTIVE: To determine the potential effects on costs and outcomes of changes in sensitivity and specificity with new screening methods for cervical cancer.

METHODS: Using a Markov model of the natural history of cervical cancer, we estimated the effects of sensitivity, specificity, and screening frequency on cost-effectiveness. Our estimates of conventional Papanicolaou test sensitivity of 51% and specificity of 97% were obtained from a meta-analysis. We estimated the effect of reducing false-negative rates from 40-90% and increasing false-positive rates by up to 20%, independently and jointly. We varied the marginal cost of improving sensitivity from $0 to $15.

RESULTS: When specificity was held constant, increasing sensitivity of the Papanicolaou test increased life expectancy and costs. When sensitivity was held constant, decreasing specificity of the Papanicolaou test increased costs, an effect that was more dramatic at more frequent intervals. Decreased specificity had a substantial effect on cost-effectiveness estimates of improved Papanicolaou test sensitivity. Most of those effects are related to the cost of evaluation and treatment of low-grade lesions.

CONCLUSION: Policies or technologies that increased sensitivity of cervical cytologic screening increased overall costs, even if the cost of the technology was identical to that of conventional Papanicolaou smears. These effects appear to be caused by relatively high prevalence of low-grade lesions and are magnified at frequent screening intervals. Efficient cervical cancer screening requires methods with greater ability to detect lesions that are most likely to become cancerous.


Characteristics of Apparently False-Negative Digene Hybrid Capture 2 High-Risk HPV DNA Testing.

Jastania R, Geddie WR, Chapman W, Boerner S.

Department of Pathology, University Health Network, and Laboratory Medicine and Pathobiology, the University of Toronto, Toronto, Canada.

Am J Clin Pathol. 2006 Feb;125(2):1-6. Abstact quote  

This study characterized cases with a negative high-risk Hybrid Capture 2 (HRHC2; Digene, Gaithersburg, MD) test result with concurrent or follow-up biopsy-confirmed high-grade cervical intraepithelial neoplasia (CIN 2/3). From 2,306 HRHC2 tests, 10 negative results were identified with CIN 2/3 (false-negative rate, 4.5%).

The majority of the patients had abnormal colposcopic findings and high-grade squamous intraepithelial lesion (HSIL) shown by concurrent cytologic examination, although with few abnormal cells. No trend was evident in the location of the dysplastic epithelium or overall lesion size. In 4 tests, the relative light units over cutoff was more than 0.4 but less than 1.0, suggesting that low quantities of human papillomavirus (HPV) DNA were present in the sample.

The negative predictive value for HRHC2 testing may be compromised when the copy number of the HPV DNA is low, and a negative HRHC2 test result may be falsely negative in patients with abnormal colposcopic findings or concurrent cytologic findings showing HSIL.
A comparison of a prototype PCR assay and hybrid capture 2 for detection of carcinogenic human papillomavirus DNA in women with equivocal or mildly abnormal papanicolaou smears.

Schiffman M, Wheeler CM, Dasgupta A, Solomon D, Castle PE; The ALTS Group.

Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Department of Health and Human Services, Bethesda, MD 20892-7234, USA.

Am J Clin Pathol. 2005 Nov;124(5):722-32. Abstract quote  

We evaluated Hybrid Capture 2 (HC2) and polymerase chain reaction (PCR) results for paired specimens collected at 19,187 visits from 5,026 of 5,060 women participating in the Atypical Squamous Cells of Undetermined Significance/Low-Grade Squamous Intraepithelial Lesion Triage Study (ALTS).

We examined the test agreement between HC2 and PCR detection for any of 13 carcinogenic human papillomavirus types targeted by HC2 and compared clinical performance of the 2 tests for detecting concurrent and follow-up cervical intraepithelial neoplasia (CIN) 3 or cancer. The k value for the 2 assays was 0.65 (95% confidence interval, 0.64-0.66), with 82.7% crude agreement. HC2 was more sensitive (93.6% vs 89.3%; P < .0005) but less specific (41.2% vs 48.5%; P < .0005) than PCR for detecting 2-year cumulative CIN 3 or cancer (n = 503). The presence of multiple types as detected by PCR and/or cytologic abnormality increased the likelihood of an HC2+ result.

Increased sensitivity of HC2 compared with PCR was surprising, given the theoretical advantages of PCR-based methods for analytic sensitivity. Smaller amounts of material used in PCR could have limited its sensitivity, but our results demonstrate the importance of optimization and standardization of PCR-based assays for clinical applications.
Reproducibility of HPV DNA Testing by Hybrid Capture 2 in a Screening Setting.

Carozzi FM, Del Mistro A, Confortini M, Sani C, Puliti D, Trevisan R, De Marco L, Tos AG, Girlando S, Palma PD, Pellegrini A, Schiboni ML, Crucitti P, Pierotti P, Vignato A, Ronco G.

Center for Study and Prevention of Cancer (CSPO), Florence, Italy
Am J Clin Pathol. 2005 Nov;124(5):716-21. Abstract quote  

Within a large Italian randomized trial on new technologies for cervical cancer screening involving 7 laboratories with different levels of experience, an intralaboratory and interlaboratory quality control program for human papillomavirus (HPV) DNA testing by Hybrid Capture 2 (HC2; Digene, Gaithersburg, MD) was implemented. To monitor the hybridization and detection steps, target samples containing purified, concentration-defined, HPV DNA were introduced in each test run.

Only 3 of 1,024 showed a mistake in a positive vs negative classification with a 1 relative light unit (RLU)/positive control specimen (PC) ratio cutoff. To monitor the preanalytic steps (particularly denaturation), blinded specimens (33 collected in PreservCyt (Cytyc, Boxborough, MA) and 36 in Specimen Transport Medium (STM, Digene) were centrally prepared, divided into aliquots, and sent to each laboratory. The multiple-rater scores for negative (<1 RLU/PC), low-positive (1 to <11 RLU/PC), and high-positive (> or =11 RLU/PC) samples, respectively, were 0.91, 0.60, and 0.69 with PreservCyt and 0.93, 0.87, and 0.90 with STM.

Our data showed high reliability and reproducibility with HC2, with values higher for STM than ThinPrep (Cytyc) samples.
Reflex High-Risk Human Papillomavirus Testing for Women With Atypical Squamous Cells of Undetermined Significance in Cytologic Smears
Effects Since Implementation in a Large Clinical Practice

Brian S. Kendall, MD, etal.
Am J Clin Pathol 2005;123:524-528 Abstract quote

Reflex high-risk human papillomavirus (HPV) testing often is used in the management of women with atypical squamous cells of undetermined significance identified in cervicovaginal screening. Following implementation of reflex testing, our laboratory processed 8,022 specimens during a 20-month period; sufficient material was available for testing in 7,334 specimens.

High-risk HPV was detected in 34.10% of these specimens. Detection rates varied with age, with positive rates as high as 58.46% in women 20 years old or younger, decreasing to 14.58% in women older than 35 years. The detection rate, categorized in 5-year age increments, showed a significant decrease until after 35 years, when the rate remained fairly constant (P < .0001). The detection rate decreased over the time of the study.

These results demonstrate that high-risk HPV detection might vary according to the age mix of the population tested and the interval after implementation of testing.
Interlaboratory Reliability of Hybrid Capture 2

Philip E. Castle, PhD, Cosette M. Wheeler, PhD, Diane Solomon, MD, Mark Schiffman, MD, and Cheri L. Peyton, for the ALTS Group*
Am J Clin Pathol 2004;122:238-245 Abstract quote

We evaluated the interlaboratory reproducibility of the Hybrid Capture 2 (HC2; Digene, Gaithersburg, MD), a test for oncogenic human papillomavirus (HPV) DNA, using data from 4 clinical center (CC) laboratories and the quality control (QC) laboratory participating in the ASCUS (atypical squamous cells of undetermined significance) and LSIL (low-grade squamous intraepithelial lesion) Triage Study (ALTS).

Residual liquid cytology specimens were tested routinely throughout the duration of ALTS at CC laboratories, and a stratified (by time in the study) random sample of specimens was retested by the HPV QC laboratory using equivalent protocols. Of the specimens selected (N = 1,175, 5.50% of all specimens obtained), 1,072 (91.23%) had sufficient specimen volume for retesting. The k value between all CC laboratories and the HPV QC laboratory was 0.84 (95% confidence interval, 0.78-0.89), with k values for individual CCs and the HPV QC laboratory ranging from 0.79 to 0.89. Agreement between test results was lowest among results for women with negative cytologic findings (0.73); among those with equivocal or abnormal cytologic findings, k values were 0.80 or more.

These data show that HC2 is a reliable test for detecting clinically relevant oncogenic HPV DNA.
Human Papillomavirus Typing With Hybrid Capture II on Archived Liquid-Based Cytologic Specimens
Is HPV Typing Always Reproducible?

Giovanni Negri, MD, Bettina Rigo, Fabio Vittadello, ScD, Eduard Egarter-Vigl, MD, and Christine Mian, ScD
Am J Clin Pathol 2004;122:90-93 Abstract quote

Reproducibility of human papillomavirus (HPV) typing on archived ThinPrep (Cytyc, Boxborough, MA) specimens was evaluated repeating Hybrid Capture II (HCII) (Digene, Gaithersburg, MD) testing after 25 to 40 months (mean, 31.3 months; group 1), 6 to 11 months (mean, 8.4 months; group 2), and 0 to 5 months (mean, 3.5 months; group 3). Another ThinPrep slide was prepared to evaluate cellularity and reproducibility of the cytologic diagnosis.

The mean residual relative light units (RLU) calculated for each group showed a strong decrease of RLU values at the second typing (group 1, 21.69%; group 2, 26.47%; and group 3, 32.25% of original values). No residual HPV DNA was shown in group 1 in 8 (13%) of 60 cases or in groups 2 and 3 in 2 (3%) of 60 cases each. These cases were associated mostly with poor cellularity and reproducibility of the initial cytologic diagnosis in the final cytologic examination. Intergroup statistical analysis of mean relative percentages for cases with satisfactory residual cellularity revealed a significant difference only between groups 1 and 3 (P < .05).

Although mostly reproducible, HPV typing results by HCII on archived specimens are influenced by material consumption. In addition, results might be affected by some DNA degradation after long-term sample storage.
High-Risk Human Papillomavirus Detection
A Split-Sample Comparison of Hybrid Capture and Chromogenic In Situ Hybridization

Carol L. Schiller, MD, etal.
Am J Clin Pathol 2004;121:537-545 Abstract quote

The American Society for Colposcopy and Cervical Pathology (ASCCP) has proposed high-risk human papillomavirus (HPV) testing as the "preferred" triage for women with atypical squamous cells of undetermined significance.

We studied 401 atypical squamous cells of undetermined significance liquid-based cervicovaginal cytology split samples for HPV by chromogenic in situ hybridization (CISH) and by Hybrid Capture (HC) II (Digene, Gaithersburg, MD); 202 underwent HC II followed by CISH, and 199 underwent CISH followed by HC II. Of 401 vials, 101 (25.2%) were positive for HPV by 1 or more methods. HC II labeled 83 of 401 (20.7%) samples as positive, while 38 of 401 (9.5%) were positive by CISH. Positive attributes of CISH include the provision of a cytomorphologic link in assessing HPV positivity and comparative ease of use in laboratories without trained molecular diagnosticians. Greater efficacy and quantitative design are advantages of HC II. Comparing data by sequence of testing showed a lower likelihood of positive test results on the second ancillary test than on the first ancillary test, regardless of age or testing method (odds ratio, second/first = 0.58; P = .003).

This finding suggests that liquid-based cervicovaginal cytology samples are not homogeneous throughout. Correlative studies with histology and polymerase chain reaction may clarify predictive values for both methods.

Can human papillomavirus DNA testing substitute for cytology in the detection of high-grade cervical lesions?

Lee KJ, Lee JK, Saw HS.

Department of Obstetrics and Gynecology, College of Medicine, Pochon CHA University, CHA General Hospital, Seoul, Korea.
Arch Pathol Lab Med. 2004 Mar;128(3):298-302. Abstract quote  

CONTEXT: High-risk human papillomaviruses (HPVs) are causal factors of cervical carcinomas.

OBJECTIVE: To evaluate the sensitivity and efficiency of HPV DNA testing in comparison with conventional cytology for detection of cervical intraepithelial neoplasia (CIN) and cancer.

DESIGN: Both testing procedures were administered to 593 women, aged 14 to 88 years (average, 41.7 years), who were referred for abnormal cytology from January 2000 through December 2001 at Korea University Guro Hospital (Seoul, Korea). After histologic confirmation by either colposcopically directed biopsy or endocervical curettage, the efficiency of the detection methods for high-grade cervical lesion was evaluated for the following 3 data sets: HPV DNA testing, conventional cytology, and the 2 tests combined.

RESULTS: The sensitivity, specificity, and positive predictive, and negative predictive values for the detection of CIN 2 or higher were 92.4%, 52.4%, 49.3%, and 93.2% for HPV DNA testing; 76.3%, 65.8%, 52.8%, and 84.7% for cytology; and 97.8%, 36.7%, 49.2%, and 97.3% for the combined tests. Among the 151 patients diagnosed with CIN 2 or CIN 3, 137 patients (90.7%) were HPV positive, 116 patients (76.8%) were proven to have abnormal cytology, and 147 patients (97.6%) were positive for either HPV DNA testing or cytology. The sensitivity values for HPV DNA testing and cytology were 97.9% (46/47) and 74.5% (35/47), respectively, for invasive cervical cancer detection, and the combined tests showed 100% (47/47) sensitivity. Depending on the patient's age and the grade of the cervical lesion, HPV DNA testing proved to be significantly more sensitive than cytology for the primary detection of cervical abnormalities (P <.001).

CONCLUSION: Human papillomavirus DNA testing for the detection of high-grade cervical lesions was more sensitive than cytology alone. In addition, the screening sensitivity can be further improved by combining cytology with HPV DNA testing. This approach is especially beneficial in detecting cancer precursors in women older than 60 years.
Efficiency of the Hybrid Capture 2 HPV DNA Test in Cervical Cancer Screening A Study by the French Society of Clinical Cytology

Patricia de Cremoux, MD, PhD, Joël Coste, MD, PhD, Xavier Sastre-Garau, MD, PhD, Martine Thioux, Christelle Bouillac, Sylvain Labbé, MD, Isabelle Cartier, MD, Marianne Ziol, MD, Anne Dosda, MD, Catherine Le Galès, PhD, Vincent Molinié, MD, Marie-Cécile Vacher-Lavenu, MD, PhD, Béatrix Cochand-Priollet, MD, PhD, Philippe Vielh, MD, PhD, and Henri Magdelénat, PhD, for the French Society of Clinical Cytology Study Group
Am J Clin Pathol 2003:120;492-499 Abstract quote

The aim of this study was to determine the efficiency of the Hybrid Capture 2 (HC2; Digene, Gaithersburg, MD) human papillomavirus (HPV) assay for the detection of cervical neoplasia. Of the 1,785 patients recruited, 462 (25.88%) were referred for colposcopy owing to previously detected cytologic abnormalities, and 1,323 (74.12%) were voluntary candidates for screening.

For all patients, a Papanicolaou smear and a monolayer smear (ThinPrep, Cytyc, Boxborough, MA) were done. HPV DNA was detected on the residual liquid-based material. False-positive results were observed in 111 cases and comprised 34 cross-reactions (1.90%) and 77 false-positive cases (4.31%) owing to a contiguous strong chemiluminescence signal. Interestingly, all these samples had a relative light units value of 1 to 3 and were contiguous to a sample with a very high HPV DNA load.

The final results showed that high-risk and low-risk HPV DNA were detected in 480 samples (26.89%) and 135 samples (7.56%), respectively. Although HC2 can be considered a reliable and sensitive test for HPV DNA detection, we do not advocate its use for large-scale screening for cervical neoplasia.

Adjunctive human papillomavirus testing in the 2-year follow-up of women with low-grade cervical cytologic abnormalities: a randomized trial and economic evaluation.

Lytwyn A, Sellors JW, Mahony JB, Daya D, Chapman W, Howard M, Roth P, Lorincz AT, Gafni A, Walter SD.

Department of Pathology, Sunnybrook and Women's College Health Sciences Centre, University of Toronto, Toronto, Ontario.
Arch Pathol Lab Med. 2003 Sep;127(9):1169-75 Abstract quote.  

CONTEXT: Although human papillomavirus (HPV) testing may aid in managing low-grade abnormality on screening cervical cytology, patient compliance with repeat testing programs requires consideration.

OBJECTIVES: To determine effectiveness and costs of repeated Papanicolaou (Pap) test and oncogenic HPV testing for detecting cervical intraepithelial neoplasia 2 or 3.

DESIGN: We conducted a randomized controlled trial of combined Pap test and cervical HPV testing by Hybrid Capture 1 test compared with Pap test alone; tests were performed every 6 months for up to 2 years. The study end point was colposcopic examination performed on all women at 2 years, or earlier if an HPV test was positive or if a Pap test showed high-grade squamous intraepithelial lesion.

SETTING: Sixty-six community family practices.

PARTICIPANTS: Two hundred fifty-seven women with atypical squamous cells of undetermined significance or low-grade squamous intraepithelial lesion on screening cervical cytology.

MAIN OUTCOME MEASURES: Detection of histologically confirmed cervical intraepithelial neoplasia 2 or 3, fully allocated costs, and loss to follow-up.

RESULTS: Combined Pap test and HPV testing detected 11 (100%) of 11 cases of cervical intraepithelial neoplasia 2/3, whereas Pap test alone detected 7 (63.6%) of these 11 cases (P =.14); corresponding specificities were 39 (46.4%) of 84 and 45 (71.4%) of 63 (P =.005). The cost-effectiveness ratio was Can $4456 per additional case of high-grade cervical intraepithelial neoplasia. Sixty-nine (26.8%) of the 257 women (24.6% combined group vs 29.1% Pap test only group, P =.41) defaulted from testing or from colposcopy when referred with an abnormal result.

CONCLUSIONS: Combined testing was more costly but may detect more cases of cervical intraepithelial neoplasia 2/3 than Pap test alone. However, poor adherence limits usefulness of a management strategy that requires repeated follow-up.

Value of human papillomavirus deoxyribonucleic acid testing after conization in the prediction of residual disease in the subsequent hysterectomy specimen.

Lin CT, Tseng CJ, Lai CH, Hsueh S, Huang KG, Huang HJ, Chao A.

Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Taoyuan, Taiwan.

Am J Obstet Gynecol 2001 Apr;184(5):940-5 Abstract quote

OBJECTIVE: Our aim was to evaluate human papillomavirus deoxyribonucleic acid testing after conization in predicting residual disease in the subsequent hysterectomy specimen.

STUDY DESIGN: A prospective study was conducted on 75 patients with grade 3 cervical intraepithelial neoplasia who had cone margins or endocervical curettage specimens showing disease and who elected to undergo hysterectomy after conization. All patients underwent high-risk human papillomavirus deoxyribonucleic acid testing by the Hybrid Capture II (Digene Corporation, Gaithersburg, MD) system before conization and at the time of hysterectomy (within 2-7 weeks after conization). The presence of human papillomavirus deoxyribonucleic acid in cells obtained by endocervical brush before hysterectomy was correlated with residual disease in the hysterectomy specimens.

RESULTS: Of the 92 patients enrolled, 75 were eligible. Of these 75 patients, 52 (69.3%) had persistent human papillomavirus deoxyribonucleic acid after conization, and 27 (36.0%) of the 75 patients had residual cervical neoplasia in the hysterectomy specimens. Those with negative results for human papillomavirus deoxyribonucleic acid after conization were all (23/23) without residual disease in the uterus (100% negative predictive value). All those who had residual disease (27/27) had positive results for human papillomavirus deoxyribonucleic acid at the time of hysterectomy (100% sensitivity). Postconization human papillomavirus deoxyribonucleic acid status (odds ratio, 4.000; 95% confidence interval, 1.531-10.449; P =.005) and grade of dysplasia after endocervical curettage (classified as grade 2 cervical intraepithelial neoplasia or less severe disease vs grade 3 cervical intraepithelial neoplasia: odds ratio, 6.612; 95% confidence interval, 2.837-15.409; P =.0002) were significantly associated with residual tumor in the uterus.

CONCLUSIONS: This prospective study confirms an excellent sensitivity and negative predictive value of human papillomavirus deoxyribonucleic acid testing after conization in predicting residual cervical neoplasia. A strategy of managing patients with grade 3 cervical intraepithelial neoplasia, based on postconization human papillomavirus deoxyribonucleic acid findings and endocervical curettage results, is proposed.

Detection of high-risk subtypes of human papillomavirus in cervical swabs: routine use of the Digene Hybrid Capture assay and polymerase chain reaction analysis.

Brennan MM, Lambkin HA, Sheehan CD, Ryan DD, O'Connor TC, Kealy WF.

Department of Histopathology, Cytology and Gynaecology, Cork University Hospital, Wilton, Cork, Ireland.

Br J Biomed Sci 2001;58(1):24-9 Abstract quote

Human papillomaviruses (HPVs) are major causative agents in the pathogenesis of cervical cancer, and more than twenty types are associated with its development. With the introduction of liquid-based preparation systems, it is envisaged that large-scale HPV testing will be established in the near future. Preliminary studies demonstrate the accessibility of these samples for DNA testing using both the Digene Hybrid Capture assay (DHCA) and polymerase chain reaction (PCR) techniques.

This study aims to assess the validity and sensitivity of the DHCA system to detect high-risk HPV DNA, using two sets of HPV consensus primers (Gp5+/Gp6+ and MY09/MY11) in tandem with routine assessment of cervical smear and biopsy samples.

Results indicate that the combination of DHCA and PCR detects more high-grade lesions than does the DHCA alone. DHCA-negative cases were categorised by subsequent PCR amplification into low-grade HPV-negative (12/16) cervical lesions and high-grade HPV-positive (7/9) cervical lesions. Gp5+/Gp6+ primers were less sensitive in detecting HPV-positive samples than was the MY09/MY11 primer set.

These results support the use of high-risk HPV testing by DHCA, with subsequent analysis of DHCA-negative samples by PCR using the MY09/MY11 primers.

Comparison of human papillomavirus DNA testing and repeat Papanicolaou test in women with low-grade cervical cytologic abnormalities: a randomized trial. HPV Effectiveness in Lowgrade Paps (HELP) Study No. 1 Group.

Lytwyn A, Sellors JW, Mahony JB, Daya D, Chapman W, Ellis N, Roth P, Lorincz AT, Gafni A.

Department of Laboratory Medicine and Pathobiology, University of Toronto, Ont.

CMAJ 2000 Sep 19;163(6):701-7 Abstract quote

BACKGROUND: Results of cervical cytology screening showing atypical squamous cells of undetermined significance (ASCUS) or low-grade squamous intraepithelial lesions (LSIL) indicate risk for high-grade cervical intraepithelial neoplasia (CIN 2 or 3). In a community-based randomized trial we compared the test performance of human papillomavirus (HPV) DNA testing with that of 6-month repeat Papanicolaou (Pap) test in detecting histologically confirmed CIN 2 or 3.

METHODS: We randomly assigned 212 women aged 16-50 years with ASCUS or LSIL on cervical cytology screening to undergo either immediate HPV DNA testing or a repeat Pap test in 6 months. Cervical swabs for the HPV DNA testing and the Pap smears were obtained by their family physicians. We tested the swabs for oncogenic HPV using the Hybrid Capture II assay (Digene Corp., Beltsville, Md.). Community-based pathologists examined the Pap smears. All women were referred for colposcopy by their family physicians. Two gynecological pathologists assessed the histology findings. We calculated test performance in women who completed the trial using CIN 2 or 3 as the reference standard.

RESULTS: A total of 159 women completed the study. Compared with HPV DNA testing, which detected 87.5% (7/8) of the cases of CIN 2 or 3, repeat Pap smear showing high-grade intraepithelial neoplasia (HSIL) detected 11.1% (1/9) of cases (p = 0.004), and repeat Pap smear showing ASCUS, LSIL or HSIL detected 55.6% (5/9) (p = 0.16). Corresponding specificities were 50.6%, 95.2% (p = 0.002) and 55.6% (p = 0.61). Loss to follow-up was 17.1% in the HPV test group and 32.7% in the repeat Pap group (p = 0.009). Given the 7 cases of CIN 2 or 3 detected by HPV testing and the 5 cases detected by the repeat Pap smear, the incremental cost of HPV testing was calculated to be $3003 per additional case of CIN identified.

INTERPRETATION: HPV DNA testing was more costly but was associated with significantly less loss to follow-up. It may detect more cases of CIN 2 or 3 in women with low-grade cytologic abnormalities.

Benefits and Costs of Using HPV Testing to Screen for Cervical Cancer.

Mandelblatt JS, Lawrence WF, Womack SM, Jacobson D, Yi B, Hwang YT, Gold K, Barter J, Shah K.

Lombardi Cancer Center, 2233 Wisconsin Ave NW, Suite 317, Washington, DC 20007.

JAMA 2002 May 8;287(18):2372-81 Abstract quote

CONTEXT: Despite quality assurance standards, Papanicolaou (Pap) test characteristics remain less than optimal.

OBJECTIVE: To compare the societal costs and benefits of human papillomavirus (HPV) testing, Pap testing, and their combination to screen for cervical cancer.

DESIGN, SETTING, AND POPULATION: A simulation model of neoplasia natural history was used to estimate the societal costs and quality-adjusted life expectancy associated with 18 different general population screening strategies: Pap plus HPV testing, Pap testing alone, and HPV testing alone every 2 or 3 years among hypothetical longitudinal cohorts of US women beginning at age 20 years and continuing to 65 years, 75 years, or death.

MAIN OUTCOME MEASURE: Discounted costs per quality-adjusted life-year (QALY) saved of each screening strategy.

RESULTS: Maximal savings in lives were achieved by screening every 2 years until death with combined HPV and Pap testing at an incremental cost of $76 183 per QALY compared with Pap testing alone every 2 years. Stopping biennial screening with HPV and Pap testing at age 75 years captures 97.8% of the benefits of lifetime screening at a cost of $70 347 per QALY. Combined biennial HPV and Pap testing to age 65 years captures 86.6% of the benefits achievable by continuing to screen until age 75 years. Human papillomavirus screening alone was equally effective as Pap testing alone at any given screening interval or age of screening cessation but was more costly and therefore was dominated. In sensitivity analyses, HPV testing would be more effective and less costly than Pap testing at a cost threshold of $5 for an HPV test.

CONCLUSIONS: Screening with HPV plus Pap tests every 2 years appears to save additional years of life at reasonable costs compared with Pap testing alone. Applying age limits to screening is a viable option to maintain benefits while reducing costs.

Cost-effectiveness of Alternative Triage Strategies for Atypical Squamous Cells of Undetermined Significance.

Kim JJ, Wright TC, Goldie SJ.

Department of Health Policy and Management, Harvard Center for Risk Analysis, 718 Huntington Ave, Second Floor, Boston, MA 02115.

JAMA 2002 May 8;287(18):2382-90 Abstract quote

CONTEXT: Every year approximately 2 million US women are diagnosed as having a cervical cytological result of atypical squamous cells of undetermined significance (ASC-US).

OBJECTIVE: To determine the most efficient and cost-effective management strategy for women in the United States diagnosed as having ASC-US.

DESIGN AND SETTING: Cost-effectiveness analysis of data from clinical trials, prospective studies, and other published literature. A computer-based model was used to compare 4 management strategies for a cytological result of ASC-US: immediate colposcopy; human papillomavirus (HPV) triage, which includes colposcopy if high-risk HPV types are detected; repeat cytology, which includes follow-up cytology at 6 and 12 months and referral for colposcopy if a repeat abnormal result occurs; and reclassifying ASC-US as normal in which a cytological result of ASC-US is ignored. Reflex HPV DNA testing uses either residual liquid-based cytological specimens or samples co-collected at the time of the initial screening for conventional cytology. Another method, referred to as the 2-visit HPV DNA triage, requires a woman with an ASC-US result to return within 1 month to provide another speciman sample.

MAIN OUTCOME MEASURES: Years of life saved (YLS), quality-adjusted life-years (QALYs), and incremental cost-effectiveness ratios. RESULTS: The least costly strategy for biennial screening was to reclassify ASC-US as normal, resulting in a reduction in total cancer incidence of 75% for conventional cytology and 84% for liquid-based cytology compared with no screening. The next least costly strategy was HPV DNA testing resulting in a reduction in total cancer incidence of 86% for conventional cytology and 90% for liquid-based cytology, followed by immediate colposcopy with a reduction of 87% and 91%, respectively. Compared with reflex HPV DNA testing, a strategy of repeat cervical cytology or delayed HPV testing costs more but is less effective. When all strategies were compared simultaneously, varying frequency and type of cytological test, biennial (vs every 3 years) liquid-based cytology with reflex HPV testing had a cost of $174 200 per YLS. In a similar comparison, liquid-based cytology with reflex HPV testing conducted every 3 years (vs every 5 years) had a cost of $59 600 per YLS and was more effective and less costly than a strategy of conventional cytology incorporating repeat cytology or immediate colposcopy conducted biennially.

CONCLUSION: Reflex HPV DNA testing provides the same or greater life expectancy benefits and is more cost-effective than other management strategies for women diagnosed as having ASC-US.

The Efficacy of Reprocessing Unsatisfactory Cervicovaginal ThinPrep Specimens With and Without Glacial Acetic Acid
Effect on Hybrid Capture II Human Papillomavirus Testing and Clinical Follow-up

S. Nicholas Agoff, MD,1 Thomas Dean, MD,2 Brigette K. Nixon, MD,1 Kathy Ingalls-Severn, SCT(ASCP),1 Linda Rinker, MT(NCA),1 and Verena S. Grieco, MD

Am J Clin Pathol 2002;118:727-732 Abstract quote

We sought to determine the efficacy of remaking initially unsatisfactory cervicovaginal ThinPrep (Cytyc, Boxborough, MA) specimens with and without the addition of glacial acetic acid (GAA) and the effect on human papilloma virus (HPV) Hybrid Capture II (HC2; Digene, Gaithersburg, MD) testing.

A total of 583 initially unsatisfactory ThinPrep slide preparations were identified, and remakes were made with the residual in the PreservCyt (Cytyc) vials with (n = 455) or without (n = 128) GAA. Clinical follow-up information was obtained. The addition of GAA resulted in a 56.5% reduction in unsatisfactory cases, compared with a 26.6% reduction without GAA. Neoplasia and atypia were detected in the reprocessed specimens. The addition of GAA resulted in false-positive HC2 test results in 10 of 10 cases. Neutralization of the specimen may reverse this effect.

Reprocessing unsatisfactory ThinPrep specimens with GAA can substantially reduce the overall unsatisfactory rate and result in the detection of significant lesions. However, the addition of GAA can result in false-positive results on HC2 HPV tests.

Test Performance Comparison of Inform HPV and Hybrid Capture 2 High-Risk HPV DNA Tests Using the SurePath Liquid-Based Pap Test as the Collection Method

Sharon Davis-Devine, CT(ASCP), etal.
Am J Clin Patholl 2005;124:24-30 Abstract quote

No US Food and Drug Administration–approved method is available to test for human papillomavirus (HPV) DNA in the SurePath liquid-based Papanicolaou (Pap) test (TriPath Care Technologies, Burlington, NC).

We compared the performance characteristics of Inform HPV (Ventana Medical Systems, Tucson, AZ) and Hybrid Capture 2 (HC2; Digene, Gaithersburg, MD) using SurePath for collection. The study included samples from 102 women, aged 18 to 60 years, who underwent a SurePath Pap test immediately before colposcopy and cervical biopsy. Each SurePath specimen was processed additionally for Inform HPV and HC2. Performance characteristics were determined by using the cervical biopsy as the "gold standard." The SurePath, Inform HPV, and HC2 correctly identified high-grade squamous intraepithelial lesions (HSILs) in 15 (94%) of 16, 6 (38%) of 16, and 14 (93%) of 15 cases, respectively.

When only SurePath samples with a diagnosis of atypical glandular and/or squamous cells were examined, Inform HPV and HC2 correctly identified HSIL in 19 (43%) of 44 and 45 (100%) of 45 cases, respectively.

When SurePath is used for collection, Inform HPV lacks sufficient sensitivity to detect HSIL reliably.

Use of the Thin Prep Pap Test in clinical practice.

Guidos BJ, Selvaggi SM.

Department of Pathology, Loyola University Medical Center, Maywood, Illinois 60153, USA.

Diagn Cytopathol 1999 Feb;20(2):70-3 Abstract quote

The Thin Prep Pap Test (Cytyc Corp., Boxborough, MA) received approval by the Food and Drug Administration in May 1996 as an alternative to the traditional conventional smear. The present direct-to-vial study assessed the utility of thin-layer technology for cervicovaginal screening in clinical practice.

From May 1997-February 1998 (10 mo), 15,006 cervical smears were processed and evaluated; of these, 5,423 (36.1%) were conventional smears (CS) and 9,583 (63.9%) were Thin Prep slides (TP). Both methods were analyzed to compare specimen adequacy and detection rates of cervical lesions. The TP method reduced the "satisfactory but limited by" rate by 97% and the unsatisfactory rate by 63%. For low-grade squamous intraepithelial lesions (LSILs), TP slides yielded 3.6% (348/9,583) as compared to 0.98% (53/5,423) for CS, an increase of 267%. The TP method detected a threefold increase in the number of high-grade squamous intraepithelial lesions (HSILs) of 1.0% (100/9,583), as compared to 0.3% (17/5,425) for the CS group. The atypical squamous cells of undetermined significance/squamous intraepithelial lesion (ASCUS SIL) ratio was reduced by 54% in the TP group.

In routine usage in our laboratory, the Thin Prep Pap Test yielded a significant increase in the detection of LSILs and HSILs as compared to conventional smears. Specimen adequacy was significantly improved.

The unsatisfactory ThinPrep Pap Test: missed opportunity for disease detection?

Bentz JS, Rowe LR, Gopez EV, Marshall CJ.

Department of Pathology, University of Utah School of Medicine, Salt Lake City 84132, USA.


Am J Clin Pathol 2002 Mar;117(3):457-63 Abstract quote

Cervical cytology specimens classified as "unsatisfactory for interpretation" represent a potential source of undetected disease.

This prospective study analyzed the potential benefits of a laboratory procedure to reprocess unsatisfactory ThinPrep Pap Tests (Cytyc, Boxborough, MA). All unsatisfactory ThinPrep samples were reprocessed using a glacial acetic acid wash. The study period unsatisfactory rate was compared with that for the previous 12 months.

The initial unsatisfactory rate was 1.3% (197/15,154). Of the unsatisfactory ThinPrep samples, 55.8% (110/197) had residual material for reprocessing. After reprocessing, 67.3% (74/110) were reclassified as "satisfactory" or "satisfactory but limited by," and the final unsatisfactory rate was 0.8% (123/15,154), a 62% decrease. Compared with the previous 12-month rate of 0.9% (209/23,730), this was a 12% reduction.

Seven (6.4%) of 110 initially classified as unsatisfactory contained an epithelial abnormality (atypical squamous cells of undetermined significance, 3; atypical glandular cells of undetermined significance, 2; low-grade squamous intraepithelial lesion, 1; squamous cell carcinoma, 1) on the reprocessed slide.

Reprocessing of unsatisfactory ThinPrep slides yielded additional cellular abnormalities that otherwise would have been undetected. The present study confirms that reprocessing of unsatisfactory ThinPrep slides is a beneficial laboratory procedure


The AutoPap 300 QC System multicenter clinical trials for use in quality control rescreening of cervical smears: I. A prospective intended use study.

Patten SF Jr, Lee JS, Wilbur DC, Bonfiglio TA, Colgan TJ, Richart RM, Cramer H, Moinuddin S.

NeoPath Inc., Redmond, Washington, USA.

Cancer 1997 Dec 25;81(6):337-42 Abstract quote

BACKGROUND: The AutoPap 300 QC System is an automated device for the analysis of conventionally prepared cervical cytology slides. The AutoPap System selects an enriched population of cases for human quality control (QC) review. The device assigns a score based on the likelihood that a slide is abnormal. Cases are selected for QC rescreening that have scores exceeding a preset threshold corresponding to approximately the top 10% (or greater) of scores.

METHODS: AutoPap false-negative detection, compared with a 10% random QC process, was tested in a 6-center clinical evaluation study. At each site, a block of up to 150 consecutive negative slides (including detected false-negative cases) were selected randomly daily. All slides were run on the AutoPap System and rescreened by cytotechnologists for truth determination. The false-negative cases included in the top 10% group selected by AutoPap System then were compared with false-negative detection by the random selection process.

RESULTS: Fourteen thousand nine hundred fourteen cases were analyzed. The AutoPap-enriched 10% quality control group contained false-negative cases at rates 3 to 5 times that of the random selection method (P < 0.01). The sensitivity for all false-negative cases was 35% and was 52% for false-negative cases at the level of low grade squamous intraepithelial lesion and higher.

CONCLUSIONS: The AutoPap 300 QC System provides the potential for a marked increase in the number of false-negative cervical cytology cases that can be detected on QC rescreening. A significant reduction in laboratory false-negative rates can be expected if this device is utilized in routine practice.

The AutoPap 300 QC System multicenter clinical trials for use in quality control rescreening of cervical smears: II. Prospective and archival sensitivity studies.

Patten SF Jr, Lee JS, Wilbur DC, Bonfiglio TA, Colgan TJ, Richart RM, Cramer H, Moinuddin S.

NeoPath, Inc., Redmond, Washington, USA.

Cancer 1997 Dec 25;81(6):343-7 Abstract quote

BACKGROUND: The AutoPap 300 QC System is an automated device for the analysis of conventional cervical cytology slides. The AutoPap selects an enriched population of cases for quality control review. These studies evaluated the overall sensitivity of AutoPap device to a wider variety of cytologic abnormalities than could be analyzed in the prospective portion of the clinical trials.

METHODS: At five clinical trial sites, positive cases were selected from: 1) archives (historic sensitivity study) or 2) current positive cases (current archive sensitivity study). Cases were analyzed by the AutoPap System along with matched negative cases and stratified into detection deciles by instrument score for each diagnostic category.

RESULTS: For the historic sensitivity study, the percentages of cases present within the top 10% instrument scores were as follows: atypical glandular cells of undetermined significance (AGUS): 33.7% (n = 243); low grade squamous intraepithelial lesion (LSIL): 57% (n = 412); high grade squamous intraepithelial lesion (HSIL): 81.6% (n = 385); and carcinoma: 77.7% (n = 139). For the current archive sensitivity study, the percentage of cases within the top 10% of instrument scores were as follows: atypical squamous cells of undetermined significance (ASCUS): 37.6% (n = 205); AGUS: 27.3% (n = 22); LSIL: 53.7% (n = 410); HSIL: 80.7% (n = 202); and carcinoma: 62.5% (n = 8). Sensitivities at higher percentiles proportionately were greater.

CONCLUSIONS: The AutoPap 300 QC System is detection sensitive, particularly at the level of LSIL and greater. When coupled with the data from the prospective intended use study, these results provided confirmation of the instrument's ability to enhance the false-negative detection rate significantly when compared with a 10% random case selection process. In addition, high sensitivities to all categories of abnormality suggest the possibility of using such instrumentation as a primary screening device for cervical cytology.

Does use of the AutoPap assisted primary screener improve cytologic diagnosis?

Bibbo M, Hawthorne C, Zimmerman B.

Department of Pathology and Cell Biology, Jefferson University Hospital, Philadelphia, Pennsylvania, USA.

Acta Cytol 1999 Jan-Feb;43(1):23-6 Abstract quote

OBJECTIVE: To correlate the ranked review slides by the AutoPap Assisted Primary Screener with the final cytologic diagnosis and to assess whether the ranking of slides improves diagnostic accuracy.

STUDY DESIGN: A total of 5,865 consecutive conventional and suitable cervical/vaginal smears, including high-risk (HR) cases, were processed by the AutoPap System. All slides designated Review were manually screened by two cytotechnologists using the Ranked Review Report. All abnormal findings and reactive/reparative changes were referred to two attending cytopathologists for a final diagnosis. After screening, the Review slides called Within Normal Limits (WNL) were selected according to rank for a further Quality Control (QC) review by the supervisor. All HR cases not selected by AutoPap for QC review were also rescreened. The final diagnoses of the possibly abnormal/reactive/reparative referred slides were recorded and distributed in five ranks according to the Ranked Review Report assignment.

RESULTS: Of 5,865 slides, 5,120 (87%) qualified for scanning. The AutoPap System designated 3,840 (75%) slides for manual review. One thousand three hundred forty-five slides were assigned for QC review. After elimination of nonqualified slides, 763 remained. Rescreening detected 1 high grade squamous intraepithelial lesion (HSIL), 2 low grade squamous intraepithelial lesions (LSIL), 5 atypical squamous cells of undetermined significance (ASCUS) and 5 Benign Reactive Changes (BRC). QC of 364 HR cases revealed 1 LSIL, 2 ASCUS and 3 BRC. Ultimately, 313 (8.1%) smears were diagnosed as ASCUS or a more severe abnormality, 262 (6.8%) as reactive/reparative changes, 3,259 (84.8%) as WNL and 6 (0.1%) as unsatisfactory. Of 313 abnormal slides, 181 were ranked by the system in the 1st rank, 61 in the 2nd, 38 in the 3rd, 19 in the 4th and 14 in the 5th. No HSIL or more severe lesions occurred in the fourth and fifth ranks.

CONCLUSION: Our study validated the claim by the manufacturer that a significant epithelial abnormality is more likely to be present if a high score is assigned to a slide. The preliminary results support use of the AutoPap Assisted Primary Screener to improve cytologic diagnosis.

Sensitivity studies of AutoPap System Location-Guided Screening of cervical-vaginal cytologic smears.

Huang TW, Lin TS, Lee JS.

Taipei Institute of Pathology, Taiwan, Republic of China.

Acta Cytol 1999 May-Jun;43(3):363-8 Abstract quote

OBJECTIVE: To present the results of a study that assessed the efficacy of a cervical cytology screening method utilizing the AutoPap System with Location-Guided Screening (AutoPap LGS) software for detecting abnormal Papanicolaou smear slides.

STUDY DESIGN: Two hundred cases of abnormal cervical and vaginal smears were selected from the recent archives of the Taipei Institute of Pathology. For each abnormal slide, a matched "within normal limit" slide was included in the study. The slides were processed on the AutoPap Primary Screening System to select slides for Review or No Review and identify areas of the Review slides for human review and diagnosis (AutoPap LGS). The effectiveness of AutoPap LGS for detecting abnormal Papanicolaou smear slides was evaluated at multiple No Review rates.

RESULTS: The AutoPap LGS demonstrated statistically superior sensitivity over current laboratory practice for the identification of abnormal slides. Assessing the potential benefit of the AutoPap LGS using a projection method, it is expected that the AutoPap LGS would detect an additional 52 low grade squamous intraepithelial lesion and 13 high grade squamous intraepithelial lesion cases missed by current laboratory practice in a population of 2,860 cases.

CONCLUSION: The effectiveness of AutoPap LGS was demonstrated by its statistically superior performance in the detection of missed abnormal slides as compared to current laboratory practice at the Taipei Institute of Pathology.

Performance of the AutoPap Primary Screening System in the detection of high-risk cases in cervicovaginal smears.

Alasio LM, Alphandery C, Grassi P, Ruggeri M, De Palo G, Pilotti S.

Department of Pathology, Istituto Nazionale Per lo Studio e la Cura dei Tumori, Milan, Italy.

Acta Cytol 2001 Sep-Oct;45(5):704-8 Abstract quote

OBJECTIVE: To assess the performance of the AutoPap Primary Screening System (APSS) (TriPath Imaging, Inc., Burlington, North Carolina, U.S.A.) for the detection of high grade cervical squamous intraepithelial lesions and invasive cervical cancer.

STUDY DESIGN: A total of 14,779 consecutive conventional Pap smears were processed by the APSS. All slides designated as "Review" by the device were manually screened according to the Bethesda System. The ranking scores obtained from the device were compared with the cytologic interpretations in all cases and with the final histologic diagnoses in the cases with cytologic severe abnormalities.

RESULTS: The device classified 10,349 slides as Review (78%) and 2,912 (22%) as "No Further Review." In the 78% Review cases, the samples were ranked in descending order of potential abnormality, broken into quintiles. The correlation between the slide quintile ranks and the manual cytologic diagnosis indicated that 90% of abnormal smears were categorized by the device as in the first and second quintile rank, and the correlation between the rank report of the device and the histologic diagnosis showed that all cases of HSIL or invasive carcinoma were in the top two ranks. No significant abnormalities were observed in any of the smears categorized as No Further Review.

CONCLUSION: This study confirmed the effectiveness of APSS for the detection of Pap smears with severe abnormalities.

Direct-to-vial experience with AutoCyte PREP in a small New England regional cytology practice.

Marino JF, Fremont-Smith M.

Seacoast Pathology P.A., 1 Hampton Road, Suite 108, Exeter, NH 03833, USA.

J Reprod Med 2001 Apr;46(4):353-8 Abstract quote

OBJECTIVE: To evaluate and assess the efficiency and efficacy of the AutoCyte PREP thin-layer Pap test method (TriPath Imaging, Inc., Burlington, North Carolina) in a privately owned cytology laboratory.

STUDY DESIGN: Data from 35,496 conventional Pap smear preparations performed in 1999 were compared to 6,357 conventional Pap smears and 15,534 AutoCyte PREP cases that were collected, prepared and processed from January 1 to July 31, 2000.

RESULTS: The AutoCyte PREP demonstrated a statistically significant increased detection of low grade squamous intraepithelial lesions (LSIL) (47%) (P = .0011) and high grade squamous intraepithelial lesions (HSIL) (116%) (P = .0002) when compared to conventional Pap smears processed during the same time period. When compared to the conventional Pap smears from 1999, the LSIL lesions increased by 57% (P < .00001), and the HSIL lesions increased by 55% (P = .0002). Both increases are statistically significant. The atypical squamous cells of undetermined significance (ASCUS)/LSIL ratio was reduced by 48% (P < .00001) using AutoCyte PREP when compared to the 1999 conventional Pap smear experience. The unsatisfactory rate was reduced by 70%. AutoCyte PREP demonstrated improved histologic correlation in HSIL cases.

CONCLUSION: AutoCyte PREP was significantly more effective than the conventional Pap smear for the detection of both LSIL and HSIL. The ASCUS/LSIL ratio and unsatisfactory cases were also significantly reduced.

The human false-negative rate of rescreening Pap tests. Measured in a two-arm prospective clinical trial.

Renshaw AA, Lezon KM, Wilbur DC.

Department of Pathology, Baptist Hospital of Miami, Miami, Florida, USA.

Cancer 2001 Apr 25;93(2):106-10 Abstract quote

BACKGROUND: Routine quality control rescreening often is used to calculate the false-negative rate (FNR) of gynecologic cytology. Theoretic analysis suggests that this is not appropriate, due to the high FNR of rescreening and the inability to actually measure it. The authors sought to determine the FNR of manual rescreening in a large, prospective, two-arm clinical trial using an analytic instrument in the evaluation.

METHODS: The results of the Autopap System Clinical Trial, encompassing 25,124 analyzed slides, were reviewed. The false-negative and false-positive rates at various thresholds were determined for routine primary screening, routine rescreening, Autopap primary screening, and Autopap rescreening by using a simple, standard methodology.

RESULTS: The FNR of routine manual rescreening at the level of atypical squamous cells of undetermined significance (ASCUS) was 73%, more than 3 times the FNR of primary screening; 11 cases were detected. The FNR of Autopap rescreening was 34%; 80 cases were detected. Routine manual rescreening decreased the laboratory FNR by less than 1%; Autopap rescreening reduced the overall laboratory FNR by 5.7%. At the same time, the false-positive rate for Autopap screening was significantly less than that of routine manual screening at the ASCUS level (4.7% vs. 5.6%; P < 0.0001). Rescreening with the Autopap system remained more sensitive than manual rescreening at the low grade squamous intraepithelial lesions threshold (FNR of 58.8% vs. 100%, respectively), although the number of cases rescreened was low.

CONCLUSIONS: Routine manual rescreening cannot be used to calculate the FNR of primary screening. Routine rescreening is an extremely ineffective method to detect error and thereby decrease a laboratory's FNR. The Autopap system is a much more effective way of detecting errors within a laboratory and reduces the laboratory's FNR by greater than 25%.

Can technology expedite the cervical cancer screening process? A Hong Kong experience using the AutoPap primary screening system with location-guided screening capability.

Chang AR, Lin WF, Chang A, Chong KS.

Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, People's Republic of China.

Am J Clin Pathol 2002 Mar;117(3):437-43 Abstract quote

We studied the usefulness of an automated screening instrument for processing Papanicolaou (Pap) smears to determine whether it could speed human examination by recording the time to screen 1,007 cervical Pap smears with an AutoPap primary screening instrument with location-guided screening (LGS) software and by conventional microscopic examination. We also assessed the accuracy of the methods to determine preparation adequacy, and we compared the diagnosis by each method.

The AutoPap with LGS satisfactorily determined the adequacy of Pap smears and identified the marked abnormal cells for human examination. An accurate diagnosis was possible when only the marked cells were examined, and this method reduced the screening time to less than half that required for conventional screening. With low-grade squamous intraepithelial and more severe lesions as a threshold, there were 37 cases in the conventional group and 29 cases in the LGS group. With atypical squamous cells of undetermined significance as the threshold, there were 111 cases in the LGS group and 93 cases in the conventional group.

The AutoPap with LGS can significantly speed the examination of Pap smears without lowering the detection rate of clinically important lesions, thus helping alleviate the cytotechnologist shortage.

The primary screening clinical trials of the TriPath AutoPap System.

Wilbur DC, Norton MK.

Cytopathology Laboratory, The Genesee Hospital, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA.

Epidemiology 2002 May;13(3 Suppl):S30-3 Abstract quote

The AutoPap Primary Screening System has been approved recently by the United States Food and Drug Administration for the initial screening and quality control of non-"high-risk" conventional cervical cytology slides.

A prospective clinical trial using the device compared its performance in terms of sensitivity and specificity for abnormality detection with the current standard of practice, which is represented by initial manual screening and 10% random quality control rescreening.

The study showed statistically superior abnormality detection sensitivity by the AutoPap System. In addition, specificity of diagnosis was also improved. Equivalent performance was noted for categorization of slide adequacy and detection of infectious organisms and benign cellular changes. Use of the AutoPap System therefore has the potential to improve the accuracy of cervical cytologic examinations as well as to make a positive impact on the productivity of cytology laboratory personnel.

Location-guided screening of liquid-based cervical cytology specimens: a potential improvement in accuracy and productivity is demonstrated in a preclinical feasibility trial.

Wilbur DC, Parker EM, Foti JA.

Cytopathology Unit, ViaHealth, Rochester, NY, USA.

Am J Clin Pathol 2002 Sep;118(3):399-407 Abstract quote

A 2-armed, masked study was performed on 1,275 AutoCyte PREP (TriPath, Burlington, NC) slides. Manual screening (current practice [CP]) was compared with automated screening with location-guided screening (LGS) using the AutoPap system with the SlideWizard 2 automated review microscopy station (TriPath).

Cytologic adjudication determined "truth" for each slide. LGS identified more abnormal cases at all interpretive levels and classified abnormalities more specifically. For high-grade squamous intraepithelial lesions and above, the overall sensitivity of appropriate triage to pathologist review was 98.4% for LGS and 91.1% for CP. The appropriate triage for all abnormal cases was 92.1% for LGS and 87.9% for CP.

The potential exists for more accurate and productive cytologic interpretation using this system, which requires no manual screening of a portion of slides and only limited review of another portion of "predotted" slides.

Human papillomavirus testing as a cytology gold standard: comparing Surinam with the Netherlands.

Wachtel MS, Boon ME, Korporaal H, Kok LP.

1St Joseph Regional Health Center, Bryan, TX, USA.
Mod Pathol. 2005 Mar;18(3):349-53. Abstract quote  

Polymerase chain reaction to detect high-risk human papillomavirus has been suggested as a gold standard for cytology. The Netherlands and Surinam were prospectively compared in regard to the proportions of Negative, Atypical Squamous Cells of Undetermined Significance, and Squamous Intraepithelial Lesion smears that had detectable high-risk human papillomavirus. For the Netherlands, 14 600 negative, 270 Atypical Squamous Cells of Undetermined Significance and 120 Squamous Intraepithelial Lesion smears were evaluated by polymerase chain reaction. For Surinam, 150 negative, 50 Atypical Squamous Cells of Undetermined Significance, and 150 Squamous Intraepithelial Lesion smears were evaluated by polymerase chain reaction.

In all, 4% of Dutch and 80% of Surinamese negative smears had detectable high-risk human papillomavirus (chi(2)=1313, P<0.00001). In total, 41.9% of Dutch and 84% of Surinamese Atypical Squamous Cells of Undetermined Significance smears had detectable high-risk human papillomavirus (chi(2)=28, P<0.00001). Totally, 67.5% of Dutch and 94% of Surinamese SIL smears had detectable high-risk human papillomavirus (chi(2)=30, P<0.00001).

The Negative: Atypical Squamous Cells of Undetermined Significance odds ratio was 0.058 for the Netherlands and 0.762 for Surinam (chi(2)(homog)=31, P<0.00001). The Negative: Squamous Intraepithelial Lesion odds ratio was 0.020 for the Netherlands and 0.255 for Surinam (chi(2)(homog)=31, P<0.00001). The Atypical Squamous Cells of Undetermined Significance: Squamous Intraepithelial Lesion odds ratio was 0.347 for the Netherlands and 0.335 for Surinam (chi(2)(homog)=0.005, P>0.75).

Human papillomavirus DNA testing may not be a suitable gold standard in general because its use would make specificity and sensitivity prevalence-dependent. A new statistic, the percent of Negative pap smears with detectable high-risk human papillomavirus, is posited, which may be important if human papillomavirus DNA testing is used clinically.

Evaluation of human papillomavirus testing in primary screening for cervical abnormalities: comparison of sensitivity, specificity, and frequency of referral.

Kulasingam SL, Hughes JP, Kiviat NB, Mao C, Weiss NS, Kuypers JM, Koutsky LA.

HPV Research Group, 1914 N 34th St, Suite 300, Seattle, WA 98103.

JAMA 2002 Oct 9;288(14):1749-57 Abstract quote

CONTEXT: Human papillomavirus (HPV) DNA testing of women having Papanicolaou (Pap) smears showing atypical squamous cells of undetermined significance (ASCUS) has clinical usefulness. Whether HPV DNA testing alone is useful in primary screening remains to be determined.

OBJECTIVE: To determine the accuracy of HPV DNA testing for detecting cervical intraepithelial neoplasia (CIN) grade 3 or cancer (the criterion standard).

DESIGN, SETTING, AND PARTICIPANTS: Between December 1997 and October 2000, 4075 women who attended Planned Parenthood clinics in Washington State were screened simultaneously using thin-layer Pap and HPV DNA testing by a polymerase chain reaction (PCR)-based method and by a liquid-based RNA-DNA hybridization capture with signal amplification assay (signal amplification). Women who were positive for high-risk HPV types, or had Pap results of ASCUS or higher, were considered to have positive screening test results and were referred for colposcopy and biopsy. Additionally, a random sample of women with negative screening test results was referred for colposcopy. Based on individual and combined thin-layer Pap, HPV PCR, and HPV signal amplification test results from the screening and the colposcopy visits, 7 colposcopy triage strategies were defined and evaluated.

MAIN OUTCOME MEASURE: Sensitivity and specificity for detecting cervical lesions graded CIN 3 or higher for each of the 7 triage strategies.

RESULTS: The estimated prevalence of CIN 3 or higher was 3.2%. The sensitivity (95% confidence interval) of thin-layer Pap (with a result of >/=ASCUS) for identifying women with CIN 3 or higher was only 61.3% (48.5%-70.9%) compared with 88.2% (78.9%-93.8%) for HPV testing by PCR and 90.8% (83.1%-95.8%) by signal amplification. Differences in specificities were also observed: 82.4% (81.8%-83.1%) for thin-layer Pap (with a result of >/=ASCUS), 78.8% (77.9%-79.7%) for PCR, and 72.6% (69.4%-75.0%) for signal amplification. Compared with referral for colposcopy of all women with ASCUS or higher, signal amplification testing of women with ASCUS and referral of those with a positive result was about as sensitive (61.3% vs 60.3%, respectively) and significantly more specific (82.4% vs 88.9%, respectively). The strategy requiring repeat positive PCR tests on 2 visits had a sensitivity of 84.2% (75.3%-91.0%) and a specificity of 86.2% (85.1%-87.3%). All tests were more specific and less sensitive in older (>/=30 years) vs younger women.

CONCLUSIONS: Testing for HPV has higher sensitivity but lower specificity than thin-layer Pap screening. In some settings, particularly where screening intervals are long or haphazard, screening for HPV DNA may be a reasonable alternative to cytology-based screening of reproductive-age women.


What are the costs of the Pap Smear to the patient and health care?


Willingness to pay for new Papanicolaou test technologies.

Raab SS, Grzybicki DM, Hart AR, Kiely S, Andrew-JaJa C, Scioscia E Jr.

Department of Pathology and Laboratory Medicine, Allegheny General Hospital, Pittsburgh, PA 15212, USA.

Am J Clin Pathol 2002 Apr;117(4):524-33 Abstract quote

The amount of money a woman is willing to pay for liquid-based cytology technology has not been measured. In the present study, 175 women answered a questionnaire asking how much they would pay to decrease their risk of dying of cervical cancer if a new (liquid-based) Papanicolaou (Pap) test was used in place of the conventional smear. When women assumed that the new Pap test reduced the risk of dying of cervical cancer from 1 in 37,000 to 1 in 50,000, the mean amount they were willing to pay was $237. If women had more than 2 children, they were willing to pay more for the new Pap test than women with 2 or fewer children.

These data indicate that liquid-based and conventional Pap tests are undervalued and that cost-effectiveness studies generally have not taken into account the preference of women for new Pap test technologies.


INTEROBSERVER VARIATION Current Bethesda 2001 Classification
Measuring the significance of field validation in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology: how good are the experts?

Renshaw AA, Wang E, Mody DR, Wilbur DC, Davey DD, Colgan TJ; Cytopathology Resource Committee, College of American Pathologists.

Department of Pathology, Baptist Hospital of Miami, Miami, Fla, USA.
Arch Pathol Lab Med. 2005 May;129(5):609-13. Abstract quote  

CONTEXT: Expert opinion is often used as a gold standard for gynecologic cytology in the evaluation of new technologies, in the legal setting, and in the validation of cases for use in educational programs and proficiency testing. However, the reliability of expert opinion alone in selecting slides of a specific cytodiagnosis that can be reproducibly and reliably identified by subsequent reviewers has not been determined.

OBJECTIVE: To assess the ability of expert opinion to select slides that are validated in subsequent reviews.

DESIGN: In the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology, each case in every cytodiagnostic category is accepted for circulation only after review by 3 expert cytopathologists. The percentage of these cases that could not be reliably and reproducibly identified by program participants for each cytodiagnostic category ("failed field validation") was determined during the duration of the program from 1989 to 2004.

RESULTS: More than 10,000 conventional smears and ThinPrep cases were selected by the expert panel for circulation. Of these selected slides, 19% of conventional smears and 15% of ThinPrep specimens failed field validation. Compared with the overall slide performance, significantly higher percentages (P < .001) of conventional smears with reference diagnoses of unsatisfactory (51.7%), repair (58%), or low-grade intraepithelial lesion (31.8%) and of ThinPrep specimens with reference diagnoses of unsatisfactory (54.5%) and repair (76.9%) failed field validation. In contrast, significantly lower percentages of conventional smears with reference diagnoses of squamous cell carcinoma (4.5%), high-grade squamous intraepithelial lesion (9%), Trichomonas vaginalis infection (11.7%), or herpes (9.9%) and of ThinPrep specimens with reference diagnoses of adenocarcinoma (5.1%), herpes (2.1%), and fungal organism consistent with Candida (8.4%) failed field validation (P < .001 for all).

CONCLUSIONS: Between 15% and 19% of gynecologic cytologic cases that have been selected by expert cytopathologists as good examples of cytodiagnostic abnormalities fail field validation. The proportion of cases failing field validation varies with cytodiagnostic category, but it occurs in all cytodiagnostic entities.

Precision in gynecologic cytologic interpretation: a study from the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology.

Renshaw AA, Davey DD, Birdsong GG, Walsh M, Styer PE, Mody DR, Colgan TJ; College of American Pathologists Comparison Program in Cervicovaginal Cytology.

Department of Pathology, Baptist Hospital, Miami, Fla, USA.

Arch Pathol Lab Med. 2003 Nov;127(11):1413-20. Abstract quote  

CONTEXT: Numerous studies address the accuracy or positive predictive value of cytologic interpretations for defined histopathologic entities. The reproducibility (precision) of cytologic interpretation is less well defined.

OBJECTIVE: To establish and compare the reproducibility (precision) of cytologic interpretation in gynecologic cytopathology, as reflected in the educational program of the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology (PAP).

METHODS: The pathologists' interpretations for both validated (25 745 responses) and educational conventional (14 353 responses) slides in the PAP program for 2001 were analyzed. The frequency of exact matches between the reference and pathologists' interpretation for each of the cytologic interpretative categories was identified, and the cumulative distributions of exact match rates were derived. chi2 Tests by reference interpretations were used for cytodiagnostic categories, least and most reproducible groupings, and high-grade (HSIL) versus low-grade (LSIL) squamous intraepithelial lesions.

RESULTS: Pathologists' interpretations of negative, Candida, Trichomonas, herpes, and LSIL were characterized by a high degree of exact matching, while interpretations of repair, HSIL, adenocarcinoma, and squamous cell carcinomas were characterized by a lesser degree of exact matching (reproducibility). Pathologists' cytologic interpretations of HSIL were significantly less reproducible than those of LSIL.

CONCLUSION: The cytologic interpretations of the most significant categories (HSIL, squamous cell carcinoma, and adenocarcinoma) are less precise than those of specific infection (Candida, Trichomonas, and yeast), negative, and LSIL categories. Cytologic interpretations of LSIL are made with greater precision than those of HSIL and may represent a more appropriate endpoint to measure the precision performance of gynecologic cytology laboratories.

Cytologic features of adenocarcinoma, not otherwise specified, in conventional smears: comparison of cases that performed poorly with those that performed well in the College of American Pathologists Interlaboratory Comparison Program in cervicovaginal cytology.

Renshaw AA, Schwartz MR, Wang E, Haja J, Hughes JH.

Department of Pathology, Baptist Hospital of Miami, Miami, FL 33176-2197, USA.

Arch Pathol Lab Med. 2006 Jan;130(1):23-6. Abstract quote  

CONTEXT: Characteristic cytologic features have been identified that distinguish cases that are consistently identified from those that are sometimes missed in the College of American Pathologists Gynecologic Cytology Program for a variety of different lesions and preparations.

OBJECTIVES: To compare the cytologic features of cases of adenocarcinoma, not otherwise specified (NOS), in conventional smears that perform poorly and well.

DESIGN: The cytologic features of 21 conventional smear cases of adenocarcinoma, NOS, that performed poorly in the College of American Pathologists Interlaboratory Comparison Program were compared with 17 cases that performed extremely well.

RESULTS: Cases that performed well were significantly more likely to have greater than 1000 abnormal cells (P = .006), greater than 100 large abnormal cells (P = .006), large nuclei (P < .001), marked nuclear atypia (P = .02), and hyperchromasia (P = .02). Repair-like features were rare in both groups and were not significant (P = .71).

CONCLUSION: Conventional smears with a diagnosis of adenocarcinoma that were consistently identified were significantly more likely to have more abnormal cells, larger abnormal cells, larger nuclei, marked atypia, and hyperchromasia than cases that performed poorly.
Atypical Glandular Cells of Undetermined Significance Outcome Predictions Based on Human Papillomavirus Testing

Jeffrey F. Krane, MD, PhD, Kenneth R. Lee, MD, Deqin Sun, MS, Liping Yuan, MD, and Christopher P. Crum, MD
Am J Clin Pathol 2004;121:87-92 Abstract quote

Cases of atypical glandular cells (AGC) diagnosed on liquid-based preparations were culled from a 3-year period. When available, residual cellular material was analyzed for human papillomavirus (HPV) by polymerase chain reaction and correlated with cytologic and histologic (biopsy) outcome.

Of 178,994 cytologic cases, 187 (0.1045%) contained AGC compared with 8,740 (4.8828%) atypical squamous cells (ASC) for an AGC/ASC ratio of 0.021. HPV results and follow-up were available for 108 specimens from 106 patients. Depending on the end-point (histologic/cyto-logic), the sensitivity range of HPV testing for significant cervical disease (high-grade squamous intraepithelial lesion [SIL], adenocarcinoma in situ [ACIS], invasive carcinoma) was 83% with a specificity range of 78% to 82%, a positive predictive value of 57% to 61%, and a negative predictive value of 91% to 95%. Fifteen false-positive results included concurrent ASC or low-grade SIL, ASC on follow-up cytology, and previous ACIS with a negative follow-up cone biopsy result.

Noncervical glandular neoplasia (including atypical endometrial hyperplasia) was confirmed in 13 cases (1 recurrent), only 2 of which scored positive for HPV.

HPV-positive AGC has a substantially higher positive predictive value for significant disease than ASC (61% vs historic 20%) and merits consideration in the triage of patients with atypical endocervical cells not otherwise specified. However, noncervical or other HPV-negative glandular neoplasia must be considered in all patients with AGC, particularly older patients.

Atypical Glandular Cells of Undetermined Significance (AGUS) Interobserver Reproducibility in Cervical Smears and Corresponding Thin-Layer Preparations

Kenneth R. Lee, MD
Teresa M. Darragh, MD
Nancy E. Joste, MD
Jeffrey F. Krane, MD, PhD
Mark E. Sherman, MD
Leo B. Hurley, MPH
Elizabeth M. Allred, MS
and M. Michele Manos, PhD, MPH

Am J Clin Pathol 2002;117:96-102 Abstract quote

Five panelists independently reviewed 135 consecutive conventional cervical smears (CPs) originally classified as atypical glandular cells of undetermined significance (AGUS).

A thin-layer slide (TP), prepared from the residual material, also was reviewed in each case. All patients underwent colposcopy that yielded at least 1 histologic specimen. Three or more of 5 reviewers retained the AGUS interpretation for 29% of CPs and 12% of the corre-sponding TPs.

Interobserver variability in frequency of use of AGUS was marked, and interobserver agreement was poor. Agreement was improved for cases cytologically interpreted as a high-grade lesion, especially in TPs. Four of 5 reviewers retained the AGUS classification in CPs for all 7 biopsy-proven neoplastic glandular lesions. Of 95 CP interpretations made by 5 reviewers in the 19 histologically diagnosed high-grade lesions, 8 were "negative/reactive" and 6 were AGUS "favor reactive." AGUS is a poorly reproducible cytologic interpretation. Although most neoplastic glandular lesions may be distinguished by cytopathologists experienced in this area from mimics originally considered AGUS, attempts to increase the diagnostic specificity of AGUS may diminish sensitivity for an underlying high-grade precursor.

Interobserver agreement was better for TPs than for the corresponding CPs. However, the split-sample TP slides may not have been fully comparable to the CPs.


Comparison of human papillomavirus DNA prevalence in atypical squamous cells of undetermined significance subcategories as defined by the original Bethesda 1991 and the new Bethesda 2001 Systems.

Pirog EC, Erroll M, Harigopal M, Centeno BA.

Department of Pathology, The Papanicolaou Laboratory, Weill Medical College of Cornell University, New York, NY 10021, USA.
Arch Pathol Lab Med. 2004 May;128(5):527-32. Abstract quote  

CONTEXT: The new Bethesda System 2001 (TBS 2001) minimized the subclassification of atypical squamous cells of undetermined significance (ASCUS).

OBJECTIVE: The primary goal of this study was to determine the impact of the new subclassification on the accuracy of Papanicolaou (Pap) test diagnosis by examining the prevalence of human papillomavirus (HPV) DNA in different ASCUS subcategories, as defined by the new TBS 2001 versus the original TBS 1991. The second goal was to identify specific morphologic features of atypical squamous cells that are more frequently associated with HPV detection.

DESIGN: Consecutive cases of ThinPrep Pap tests were retrospectively reviewed by a panel of pathologists to obtain consensus diagnoses. The study group consisted of ASCUS cases; the positive control group consisted of low- and high-grade squamous intraepithelial lesions (LSILs and HSILs, respectively); and the negative control group consisted of cases "negative for intraepithelial lesion or malignancy." All ASCUS cases were subclassified according to TBS 1991 into the following categories: favor reactive (ASCUS-R), favor LSIL (ASCUS-L), favor HSIL (ASCUS-H), and not otherwise specified (ASCUS-NOS). In a separate review, ASCUS cases were subclassified according to TBS 2001 into the following categories: atypical squamous cells of undetermined significance (ASC-US) and atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion (ASC-H). Furthermore, morphologic ASCUS subtypes were recorded (atypical mature, immature, parakeratotic, and atrophic cells); in addition, individual morphologic features of atypical cells were recorded. Broad- spectrum HPV DNA amplification and genotyping was performed using short PCR fragment (SPF 10) polymerase chain reaction/Line Probe assays.

RESULTS: In cases classified according to TBS 1991, HPV was detected in 32% of negative, 49% of ASCUS, and 93% of LSIL/HSIL cases. On the second review, using the diagnostic categories of TBS 2001, which eliminated the ASCUS-;R category, the number of ASCUS cases decreased by 45%. The prevalence of HPV DNA in ASCUS cases downgraded to the negative category was 38%, which was not significantly different from HPV prevalence in negative cases as diagnosed under TBS 1991. Furthermore, HPV was detected in 56% of ASC-US and 71% of ASC-H cases. The prevalence of HPV in different morphologic subtypes of ASCUS was not significantly different, and none of the 8 individual morphologic features of atypical cells were more frequently associated with HPV detection.

CONCLUSION: Elimination of the ASCUS-R category in TBS 2001 resulted in a significant decrease in the number of ASCUS diagnoses. Downgraded cases had a relatively low prevalence of HPV DNA. It is expected that TBS 2001 will increase specificity of the Pap test without compromising its sensitivity.

An Analysis of Factors That Influence the ASCUS/SIL Ratio of Pathologists

Ridas Juskevicius, MD,
Kelly H. Zou, PhD,
and Edmund S. Cibas, MD

Am J Clin Pathol 2001;116:331-335 Abstract quote

In pursuit of physician-specific performance data in cytology, we have been calculating the ASCUS/SIL (atypical squamous cells of undetermined significance/squamous intraepithelial lesion) ratio of cytopathologists (CPs) and providing confidential feedback every 6 months. At the same time, thin-layer technology was introduced as an alternative to conventional smears.

Thus we analyzed factors that may influence the ASCUS/SIL ratio, particularly the effect of periodic feedback on outliers (defined by a professional benchmark). For 3 years, the mean ASCUS/SIL ratio for all CPs decreased significantly from 2.92 to 1.87. There was great variability in the mean ASCUS/SIL ratio among 12 CPs (range, 1.11-5.89). Of the 6 CPs who worked continuously during this time, 2 showed a statistically significant decrease in their ASCUS/SIL ratio, including the CP with the highest ratio; 1 showed a significant increase. The mean ASCUS/SIL ratio did not correlate well with years of CP experience or with individual annual case volume.

The ASCUS/SIL ratio of some CPs can decrease significantly over time. Whether it was due to feedback or the introduction of thin-layer preparations could not be determined.

Qualification of ASCUS A Comparison of Equivocal LSIL and Equivocal HSIL Cervical Cytology in the ASCUS LSIL Triage Study

Mark E. Sherman, MD
Diane Solomon
Mark Schiffman for the ALTS Group*

Am J Clin Pathol 2001;116:386-394 Abstract quote

Cytologic detection of high-grade squamous intraepithelial lesions (HSILs) is critical to cervical cancer prevention. Therefore, identifying "equivocal HSIL" (ASCUS [atypical squamous cells of undetermined significance]-H) may be useful. Accordingly, we compared findings associated with "equivocal low-grade SIL" (ASCUS-L), ASCUS-H, and HSIL using data from the ASCUS LSIL (low-grade squamous intraepithelial lesion) Triage Study.

The frequency of oncogenic human papillomavirus (HPV) DNA detection and underlying lesions cervical intraepithelial neoplasia (CIN) 2 or worse or CIN 3 or worse in women with ASCUS-H was intermediate between that of ASCUS-L and HSIL. Oncogenic HPV DNA was associated with 85.6% of ASCUS-H ThinPreps and 69.8% of ASCUS-H smears. Histopathologic lesions CIN 2 or worse were associated with 40.5% of ASCUS-H ThinPreps and 27.2% of ASCUS-H smears (mostly CIN 3). Nevertheless, numerically more lesions CIN 2 or worse were preceded by ASCUS-L than by ASCUS-H because ASCUS-L was more common.

ASCUS-H is an uncommon interpretation that derives clinical usefulness from its high positive predictive value for lesions CIN 2 or worse.

Should Atypical Squamous Cells of Undetermined Significance (ASCUS) Be Subcategorized? Accuracy Analysis of Papanicolaou Smears Using Receiver Operating Characteristic Curves and Implications for the ASCUS/Squamous Intraepithelial Lesion Ratio

Andrew A. Renshaw, MD
David R. Genest, MD
Edmund S. Cibas, MD

Am J Clin Pathol 2001;116:692-695 Abstract quote

We correlated all Papanicolaou test diagnoses over a 6-month period with biopsy results and determined accuracy using receiver operating characteristic curves and biopsy as the "gold standard." A

ccuracies were calculated using all atypical squamous cells of undetermined significance (ASCUS) cases or by eliminating subsets thereof. Retaining the ASCUS category resulted in significantly greater accuracy for the diagnosis of squamous intraepithelial lesion (SIL) on biopsy compared with eliminating it by diagnosing all such cases as negative.

Subcategorization significantly improved the accuracy of the test only when all cases were included. The highest accuracy without subcategorization was achieved when ASCUS, favor reactive, cases were diagnosed as negative, but this threshold was significantly less sensitive than including all ASCUS cases. Increasing or decreasing the estimated ASCUS/SIL ratio from 2.4 without subcategorization significantly reduced accuracy. Similar results were obtained when high-grade SIL on biopsy was used as the gold standard.

Use of the ASCUS category significantly improves the accuracy of the Papanicolaou test. Eliminating any subset of ASCUS reduces the ASCUS/SIL ratio but also significantly diminishes the sensitivity of the Papanicolaou test.



Clinical significance of cytologic diagnosis of atypical squamous cells, cannot exclude high grade, in perimenopausal and postmenopausal women.

Department of Pathology, Allegheny General Hospital and Drexel University College of Medicine, Pittsburgh, PA 15212, USA.


Am J Clin Pathol. 2006 Sep;126(3):381-8 Abstract quote

We used cytohistologic correlation to determine the clinical significance of atypical squamous cells, cannot exclude high grade (ASC-H) in perimenopausal and postmenopausal women.

A computer search identified 250 Papanicolaou smears from women older than 45 years with a diagnosis of ASC- H. Cases were considered perimenopausal (45 to < 55 years; 150 cases) and postmenopausal ((3)55 years; 100 cases). No follow-up data were available for 33 cases, which were excluded. The remaining 217 cases (perimenopausal, 127; postmenopausal, 90) had surgical or cytologic follow-up. Results of follow-up colposcopic biopsy were available for 176 (81.1%) and cytology for 41 (18.9%) women.

Follow-up results were as follows: perimenopausal women, negative, 50 (39.4%); mild dysplasia (low-grade squamous intraepithelial lesion [LSIL]), 46 (36.2%); high-grade dysplasia (high-grade SIL [HSIL]); 28 (22.0%); and ASC of undetermined significance (ASC-US), 3 (2.4%); postmenopausal women, negative, 52 (58%); LSIL, 31 (34%); HSIL, 5 (6%); and ASC-US, 2 (2%).

The diagnosis of ASC-H in postmenopausal women usually is associated with LSIL or a negative diagnosis on follow-up, suggesting a less aggressive surveillance and treatment regimen is needed for postmenopausal women with ASC-H.
Atypical Squamous Cells, Cannot Exclude High-Grade Squamous Intraepithelial Lesion A Follow-up Study of Conventional and Liquid-Based Preparations in a High-Risk Population

Ana Paula Louro, MD, Janie Roberson, SCT(ASCP), Isam Eltoum, MD, and David C. Chhieng, MD
Am J Clin Pathol 2003;120:392-397 Abstract quote

We compared the histologic follow-up of 368 smears or slides with an interpretation of "atypical squamous cells, cannot rule out high-grade squamous intraepithelial lesion" (ASC-H) based on conventional and liquid-based preparations and age groups in a high-risk population.

Patients with an ASC-H interpretation were 17 to 87 years old (mean, 36.8 years). The specimens were 52 liquid-based preparations and 316 conventional smears. For 218 cases (59.2%), including 28 liquid-based preparations (65%) and 190 conventional smears (58%), histologic follow-up was available. In 20 liquid-based preparations (71%) and 152 conventional smears (80.0%), cervical intraepithelial neoplasia (CIN) or higher was revealed on subsequent biopsy. Other results were as follows: liquid-based preparations, CIN1, 11 (55%); CIN2/3, 9 (45%); conventional smears, CIN1, 78 (51.3%); CIN2/3, 70 (46.1%); squamous cell carcinoma, 4 (2.6%). There was no statistically significant difference in the incidence of CIN or higher on subsequent biopsy after an interpretation of ASC-H based on preparation types.

The incidences of CIN in patients 40 years old or older and patients younger than 40 years were 66% and 84%, respectively, a statistically significant difference. Because of the high incidence of clinically significant lesions noted on subsequent follow-up, patients with an interpretation of ASC-H should be observed closely and referred for colposcopic examination regardless of their age.

Papanicolaou tests with mixed high-grade and low-grade squamous intraepithelial lesion features: distinct performance in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytopathology.

Renshaw AA, Mody DR, Styer P, Schwartz M, Ducatman B, Colgan TJ.

Department of Pathology, Baptist Hospital of Miami, Miami, FL, USA.

Arch Pathol Lab Med. 2006 Apr;130(4):456-9. Abstract quote  

CONTEXT: Previous studies have shown that in gynecologic cytology, cases of low-grade squamous intraepithelial lesion (LSIL) and high-grade squamous intraepithelial lesion (HSIL) perform differently on interpretive review. The performance of cases with mixed LSIL and HSIL features is unknown.

OBJECTIVE: To compare the performance of gynecologic cytology cases of "pure" LSIL and HSIL with cases showing mixed LSIL and HSIL features.

DESIGN: We compiled performance data from the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytopathology from the years 2003 and 2004, and compared the performance of slides showing relatively pure LSIL and HSIL (< or = 10% misclassification as HSIL and LSIL, respectively) with slides showing mixed LSIL or HSIL features (cases misclassified as LSIL or HSIL > 10% of the time).

RESULTS: Interpretations from a total of 4508 cases (2452 HSIL and 2056 LSIL) were analyzed. Overall, the sensitivity of participants on slides with a reference diagnosis of HSIL was 97.3%, and of LSIL was 95.9%. Performance trends for pure versus mixed cases varied by slide type and reference diagnosis. For conventional slides, participant sensitivity on pure HSIL cases was greatest (98.0%) and on pure LSIL cases was least (95.2%), while participant performance on cases with mixed features was intermediate (97.0% for mixed HSIL and 96.7% for mixed LSIL). In contrast, participant performance on ThinPrep slides showed the greatest sensitivity for mixed LSIL slides (97.9%), while performance on mixed HSIL slides showed the lowest sensitivity (95.7%); slides with pure features had intermediate sensitivity levels (96.3% for both HSIL and LSIL). Further evaluation demonstrated that conventional pure HSIL slides performed significantly better than mixed HSIL slides (P = .006), whereas mixed LSIL slides performed better than pure LSIL slides (P = .01). For ThinPrep slides, pure HSIL cases performed similarly to mixed HSIL cases (P = .43), while mixed LSIL cases performed better than pure LSIL cases (P = .04).

CONCLUSION: Slides with mixed LSIL and HSIL features have measurably distinct performance characteristics in comparison to slides with pure LSIL or HSIL features. Participant performance on conventional mixed cases is distinctly different from performance on ThinPrep mixed cases.
Cytologic features of high-grade squamous intraepithelial lesion in conventional slides: what is the difference between cases that perform well and those that perform poorly?

Renshaw AA, Prey MU, Hodes L, Weisson M, Haja J, Moriarty AT; Gynecologic Cytology Committee, College of American Pathologists.

Department of Pathology, Baptist Hospital of Miami, Miami, Fla 33176-2197, USA.
Arch Pathol Lab Med. 2005 Jun;129(6):733-5. Abstract quote  

CONTEXT: Previous studies have suggested that cases of high-grade squamous intraepithelial lesion in conventional smears and in ThinPrep specimens that are frequently misinterpreted as normal have relatively few small and hypochromatic dysplastic cells.

OBJECTIVE: To determine the cytologic differences between conventional Papanicolaou slides of high-grade squamous intraepithelial lesion that perform poorly and those that perform well.

DESIGN: We compared the cytologic features of 22 cases of conventional smears with high-grade squamous intraepithelial lesion that performed poorly in the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytology with 45 cases of conventional smears that performed extremely well.

RESULTS: Cases that performed poorly were significantly more likely to have 50 or fewer single dysplastic cells (P = .003) and to have only small dysplastic cells (P = .01). Cases that performed well were also more likely to have more than 500 dysplastic cells (P = .002), to exhibit the presence of large dysplastic cells (P < .001), and to be keratinized (P = .03). Hypochromasia and the number of groups of dysplastic cells were not correlated with performance.

CONCLUSIONS: Conventional smears with high-grade squamous intraepithelial lesion with 50 or fewer single dysplastic cells, no large dysplastic cells, and lacking keratinization are highly associated with poor performance in this program.
Cytologic features of high-grade squamous intraepithelial lesion in ThinPrep Papanicolaou test slides: comparison of cases that performed poorly with those that performed well in the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology.

Renshaw AA, Schulte MA, Plott E, Dubray-Benstein B, Cobb CJ, Lozano RL, Neal MH, Hughes JH, Young NA, Prey M.

Department of Pathology, Baptist Hospital of Miami, Miami, Fla 33176, USA
Arch Pathol Lab Med. 2004 Jul;128(7):746-8. Abstract quote  

CONTEXT: Conventional Papanicolaou (Pap) test slides of high-grade squamous intraepithelial lesions (HSILs) that are frequently misdiagnosed are known to have relatively few dysplastic cells. Whether this is true of cases of HSIL in ThinPrep Pap Test specimens is not known.

OBJECTIVE: To determine if cases of HSIL in ThinPrep specimens that are frequently missed have relatively few dysplastic cells.

DESIGN: The cytologic features of 16 ThinPrep cases of HSIL that performed poorly in the College of American Pathologists Interlaboratory Comparison Program were compared with 22 ThinPrep Pap Test cases that performed extremely well.

RESULTS: Significantly more cases that performed poorly had fewer than 250 dysplastic cells (13/16) than cases that performed well (3/22) (P <.001).

CONCLUSION: ThinPrep Pap Test cases with a diagnosis of HSIL that performed poorly in this program had significantly fewer dysplastic cells than those that performed well.
"Low-Grade Squamous Intraepithelial Lesion, Cannot Exclude High-Grade Squamous Intraepithelial Lesion" Is a Distinct Cytologic Category: Histologic Outcomes and HPV Prevalence.

The Department of Pathology, The Johns Hopkins Hospital, Baltimore, MD.


Am J Clin Pathol. 2007 Sep;128(3):398-403. Abstract quote

We examined the histologic outcomes and prevalence of high-risk human papillomavirus (HR-HPV) in women with liquid-based Papanicolaou (Pap) tests interpreted as "low-grade squamous intraepithelial lesion, cannot exclude high-grade squamous intraepithelial lesion" (LSIL-H) compared with the 2001 Bethesda System (TBS 2001) cytologic categories of LSIL, high-grade SIL (HSIL), and atypical squamous cells, cannot exclude HSIL (ASC-H).

A computer search identified 426 LSIL, 86 ASC-H, 81 LSIL-H, and 110 HSIL cytologic interpretations during a 1-year period, each with up to 2 years of histologic follow-up. The risk of histologic cervical intraepithelial neoplasia (CIN) 2 or worse (CIN 2+) associated with LSIL-H (32/81 [40%]) was intermediate between LSIL (46/426 [10.8%]) and HSIL (72/110 [65.5%]), but not significantly different from ASC-H (23/86 [27%]). However, LSIL-H was more frequently associated with a definitive histologic diagnosis of any CIN (CIN 1+) than ASC-H (53/81 [65%] vs 35/86 [41%]). Moreover, the prevalence of HR-HPV was significantly greater in patients with LSIL-H than in patients with ASC-H (15/15 [100%] vs 43/73 [59%]).

The histologic outcomes and HR-HPV prevalence associated with LSIL-H differ significantly from the established categories of TBS 2001 and provide evidence to support the recognition of LSIL-H as a distinct cytologic category.
Altered recognition of reparative changes in ThinPrep specimens in the College of American Pathologists Gynecologic Cytology Program.

Snyder TM, Renshaw AA, Styer PE, Mody DR, Colgan TJ; Cytopathology Resource Committee, College of American Pathologists.

Department of Pathology, The Methodist Hospital, Houston, Tex, USA.

Arch Pathol Lab Med. 2005 Jul;129(7):861-5. Abstract quote  

CONTEXT: Previous studies have shown that the diagnosis of reparative changes in conventional smears in the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytology is one of the least reproducible diagnoses. Indeed, the diagnosis of reparative changes consistently yields the highest false-positive rate of any negative for intraepithelial lesions and malignancy (NILM) cytodiagnostic category. It is unknown whether cytologists recognize reparative changes in ThinPrep specimens as well, or less often, as in conventional smears.

OBJECTIVE: To assess and compare the ability of cytologists to recognize reparative changes in conventional and ThinPrep preparations.

DESIGN: We compiled performance data from the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytology from the 2000-2003 program years. More than 400 slides with a reference diagnosis of reparative changes met our study criteria, representing a total of 11 200 individual responses for conventional cases and 1155 individual responses for ThinPrep specimens. We evaluated the results of both individual and laboratory participants using 2 performance criteria: the false-positive discordancy rate and the exact match error rate (any response that does not exactly match the reference diagnosis of 120 [reparative changes]).

RESULTS: Cases with a reference diagnosis of reparative changes made up 1.2% of all ThinPrep slides and 3.7% of all conventional slides in circulation. The false-positive discordancy rate of individual responses on educational slides for conventional smears was significantly higher than the corresponding false-positive discordancy rate for ThinPrep specimens (15.7% for conventional vs 7.1% for ThinPrep specimens, P < .001). Laboratory responses on educational conventional smears and ThinPrep slides showed a similar trend (14.2% for conventional smears vs 2.4% for ThinPrep slides, P = .002). The exact match error rate on educational conventional slides was 41.4% for individual responses, while on educational ThinPrep slides, the overall error rate was 57.5% (P < .001). For laboratory responses, the exact match error rate was 40.5% for educational conventional smears versus 58.9% for educational ThinPrep smears (P < .001). Characteristic features of reparative changes were identified in ThinPrep specimens.

CONCLUSIONS: In the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytology, ThinPrep slides with a reference diagnosis of reparative changes have a lower false-positive discordancy rate than conventional slides. Responses to ThinPrep cases with a reference diagnosis of reparative change show a higher exact match error rate than conventional smears. Since reparative changes in gynecologic cytology are recognized as indicating an increased risk of significant lesions, the clinical significance of these altered patterns of recognition of reparative changes in ThinPrep specimens warrants further investigation.
Cytologic features of squamous cell carcinoma in conventional smears: comparison of cases that performed poorly with those that performed well in the College of American Pathologists Interlaboratory Comparison Program in cervicovaginal cytology.

Renshaw AA, Henry MR, Birdsong GG, Wang E, Haja J, Hughes JH.

Department of Pathology, Baptist Hospital of Miami, Miami, FL 33176-2197, USA.
Arch Pathol Lab Med. 2005 Sep;129(9):1097-9. Abstract quote  

CONTEXT: Characteristic cytologic features have been identified that distinguish cases that are always identified from those that are sometimes missed in the College of American Pathologists Gynecologic Cytology Program for a variety of different lesions and preparations.

OBJECTIVE: To compare the cytologic features of cases of squamous cell carcinoma in conventional smears that perform poorly and well.

DESIGN: The cytologic features of 8 conventional smear cases of squamous cell carcinoma that performed poorly in the College of American Pathologists Interlaboratory Comparison Program were compared with 17 cases that performed extremely well.

RESULTS: A total of 2387 individual interpretations were recorded. Of the 86 incorrect responses, 6.2% were for repair, and 0.8% were for Trichomonas. Cases that performed well were significantly more likely to have greater than 1000 dysplastic cells (16/17 vs 4/8, P = .02) and be keratinized (13/17 vs 1/8, P = .007). Obscuring inflammation and cell size were not significant.

CONCLUSION: Conventional smears with a diagnosis of squamous cell carcinoma that were always identified were significantly more likely to have greater than 1000 cells and be keratinized than cases that performed poorly.


The Clinical Significance of Benign-Appearing Endometrial Cells on a Papanicolaou Test in Women 40 Years or Older

Tara-Jane Browne, MRCPath, David R. Genest, MD, and Edmund S. Cibas, MD
Am J Clin Pathol 2005;124:834-837 Abstract quote
The 2001 Bethesda System revision recommends reporting benign-appearing endometrial cells (BAEMC) in all women 40 years or older, not just in postmenopausal women.

We studied the influence of this change on clinical practice and the detection of endometrial neoplasia. Women 40 years or older were selected for study if BAEMC were reported on a cervical cytologic specimen. Cases were stratified by implementation of the 2001 guidelines. Results of endometrial biopsies within 18 months, with special attention to the frequency of adenocarcinoma and hyperplasia, were assessed. In the preimplementation group, BAEMC were reported as an epithelial cell abnormality in 154 postmenopausal women and as an incidental finding in 636 premenopausal women (total = 790). In the postimplementation group, BAEMC were reported in 836 women.

The proportion of endometrial biopsies in these cohorts was significantly increased (P < .05). After implementation of the 2001 guidelines, the detection of endometrial cancer increased (6 vs 3 cases, not significant). Of the women with cancer, 3 were premenopausal, 5 were asymptomatic, and 2 were premenopausal and asymptomatic.

All cancers were identified in women older than 45 years, suggesting that the Bethesda cutoff of 40 years is safe and conservative.
The Significance of Benign Endometrial Cells in Cervicovaginal Smears.

Fadare O, Ghofrani M, Chacho MS, Parkash V.

From the *Department of Pathology, Yale University School of Medicine and Yale-New Haven Hospital, New Haven, Connecticut; and daggerDepartment of Pathology, Hospital of St. Raphael, New Haven, Connecticut.

Adv Anat Pathol. 2005 Sep;12(5):274-287. Abstract quote  

The success of the routine Papanicolaou (pap) smear in reducing the incidence and mortality of cervical cancer has been chronicled extensively. Unfortunately, endometrial cancer, the most common malignancy of the gynecologic tract, continues to lack a screening modality of comparable efficacy.

It is generally accepted that the Papanicolau test has a low sensitivity for detecting endometrial disease. Nonetheless, it remains true that endometrial cells are not uncommonly identified on routine cervicovaginal smears and along with each case comes an associated responsibility for pathologists to assess cytologic features, assign a potential clinical significance, and make a decision on reporting this finding.

When endometrial cells with an entirely normal cytologic appearance are identified on an otherwise unremarkable cervicovaginal smear, the central question raised is whether such cells are exfoliating physiologically or whether their exfoliation is pathologic in response to an underlying endometrial disease. Additionally, in the former scenario, could subsets of patients be defined in which the reporting of this finding is deemed unnecessary in the cytologic report?

In this contribution, we explore the clinical significance of finding normal endometrial cells in cervicovaginal smears based on a review of the medical literature of the last half-century. The historical and evidentiary basis for the Bethesda 2001 recommendations, which calls for the reporting of cytologically benign endometrial cells only in patients 40 years and older, is reviewed in detail.
Reporting Endometrial Cells in Women 40 Years and Older
Assessing the Clinical Usefulness of Bethesda 2001

Aylin Simsir, MD, etal..
Am J Clin Pathol 2005;123:571-575 Abstract quote

We assessed the usefulness of revised Bethesda System reporting of exfoliated benign endometrial cells (EMs) in postmenopausal women. Cervicovaginal cytology specimens with benign EMs in postmenopausal women and "out-of-phase EMs" in premenopausal women 40 years and older were identified. Cases with histologic follow-up within 12 months were selected.

There was tissue follow-up for 130 postmenopausal women: 10 (7.7%) had significant findings (endometrial adenocarcinoma, 6 [2 (33%) in asymptomatic women]; complex atypical endometrial hyperplasia [CAH], 3; leiomyosarcoma, 1); 20 were receiving hormone replacement therapy (HRT; n = 15) or tamoxifen (n = 5); 2 (10%) had significant pathology (endometrial adenocarcinoma, 1; CAH, 1). Eight not taking hormones (7.3%) had significant pathology (adenocarcinoma, 5; CAH, 2; leiomyosarcoma, 1). There were follow-up data for 96 premenopausal women; only 1 (who had vaginal bleeding) had significant pathology (CAH).

The difference in incidence of preneoplastic and neoplastic conditions after a cytologic interpretation of "benign EM" between postmenopausal and premeno-pausal women was significant (P £ .025); There was no difference between postmenopausal women receiving or not receiving HRT (P > .05).

Reporting benign EMs for premenopausal women 40 years and older has no clinical significance but does for postmenopausal women, regardless of HRT and symptoms.
HYPERCHROMATIC CROWDED GROUPS This is perhaps the most difficult diagnostic area for the cytopathologist. There are several entities, both benign and malignant, which must be distinguished from each other
Changes associated with IUD
DDX with adenocarcinoma - lack of malignant features
DDX with CIS - actinomyces infection may exhibit cellular changes suggestive of CIS

Few atypical cells
Bland dark chromatin
No nucleoli
No multinucleation
Tubal ciliated metaplasia
Nuclei less crowded
Finer chromatin pattern
Clear cytoplasmic vacuoles
Atypical repair
Sheet- like arrangement “school of fish”
Finely granular chromatin prominent nucleoli
Cyanophilic vacuolated cytoplasm
Bi- and multinucleation mitosis
Exfoliated lower segment endometrium
Resembles AIS
Uniform small cells in sheets or tubules
Lack of feathering or rosettes
Fine, even chromatin
Spindled stromal cells attached
May have mitotic figures
Menstrual endometrial cells
Resembles AIS
Poorly preserved cells
Hyperchromasia of degeneration
Stromal balls and histiocytes
Absence of feathering and rosettes
Reactive endocervical cells
Spectrum of atypia
Fine or smudged chromatin
Smooth nuclear membranes
Prominent to absent nucleoli
Low N/C ratio
Minimal overlap “ lay flat”
Often associated with SIL
Microglandular hyperplasia
Endocervical adenocarcinoma in situ (AIS)

Small endometrioid type AIS may be mistaken for menstrual or lower uterine segment
Feathering, rosettes, and columnar shapes
Absence of endometrial tubules, sheets, or stroma

May also mimic reactive endocervical cells, distinguish by:
Uniform population ( not a spectrum)
Crowding, nuclear enlargement, increased N/C ratio
Coarse chromatin
Small to prominent nucleoli in every cell

No diathesis
Apoptotic bodies (70%)
Mitotic figures in 40 - 60% of cases

Invasive adenocarcinoma
Tumor diathesis
Increased balls of tumor cells
(Hi grade squamous intraepithelial lesion)
Endometrial adenocarcinoma
Cellularity - less
Cells - small,round, plump,degenerated
May have spindling of cells
Characteristic palisading indicative of glandular differentiation with a luminal surface
Groups - balls, molded groups
Cytoplasm - Vacuolated, basophilic, often with PMNs
Nuclei - small, < 2.5x intermediate cell, less hyperchromatic with single nucleolus
Squamous cell carcinoma
Atrophic parabasal cells
Fallopian tube cells
Goblet-like cells common in atrophy
Vaginal adenosis
Mesonephric remnants
Bartholin glands
Rectovaginal fistula

Significance of histiocytes in cervical smears from peri/postmenopausal women.

Tambouret R, Bell DA, Centeno BA.

James Homer Wright Laboratories, Massachusetts General Hospital, Department of Pathology, Harvard Medical School, Boston, Massachusetts 02114, USA.

Diagn Cytopathol 2001 Apr;24(4):271-5 Abstract quote

Histiocytes in cervicovaginal smears from peri- or postmenopausal women have been viewed as a potential indicator of endometrial neoplasia, but recent studies have refuted that view.

This study further defines the clinical significance of such findings. All cervical smears were selected from women (< or = 50 yr) in whom the presence of histiocytes was mentioned; pertinent clinical history, follow-up information, and selected slides were reviewed.

Among 105,225 total cervicovaginal smears from a 3.5-yr period, 106 smears from 103 women were identified. Forty-two patients (41%) were on hormone replacement therapy (2 on tamoxifen), and 23 (22.3%) patients experienced vaginal bleeding, all of whom had biopsy or cytology follow-up. Ten (9.6%) patients had no follow-up, 35 (32%) had repeat smears only, and 58 (56.3%) had endometrial/endocervical sampling. In 28 patients, the index smear was categorized as other than within normal limits or benign cellular changes; of these, 24 had subsequent tissue sampling and 4 had cytology follow-up. Of the patients with tissue sampling, 51 (84%) had benign findings (including polyps), 2 (3.5%) had hyperplasia, and 5 (10%) had adenocarcinoma. All 5 patients with adenocarcinoma had endometrial glandular cells on the smear, and 4 had vaginal bleeding. One patient with hyperplasia had vaginal bleeding, and the other was on tamoxifen and had endometrial glandular cells on the smear. None of the patients having only histiocytes on their smears and no clinical symptoms or risk factors had endometrial adenocarcinoma or hyperplasia.

These findings support the conclusion that the presence of histiocytes alone on cervicovaginal smears from peri- or postmenopausal women is nonspecific and of no major clinical significance in the absence of other clinical or cytologic findings.

Enlarged Squamous Cell Nuclei in Cervical Cytologic Specimens From Perimenopausal Women ("PM Cells")
A Cause of ASC Overdiagnosis

Edmund S. Cibas, MD, Tara-Jane Browne, MB, BCh, BAO, Michelle H. Mantel Bassichis, MD, and Kenneth R. Lee, MD
Am J Clin Pathol 2005;124:58-61 Abstract quote

We studied the appropriateness of interpreting squamous cells with enlarged, smooth, bland nuclei in perimenopausal women ("PM cells") as atypical squamous cells (ASCs). Papanicolaou smears (Paps) from 100 women (40-55 years old) with a cytologic interpretation of ASC of undetermined significance (ASCUS) and human papillomavirus (HPV) testing or a biopsy within 6 months were reviewed by 2 observers without knowledge of the biopsy diagnosis or HPV results.

Cases in which both reviewers agreed that the Paps were diagnosed more properly as "negative for intraepithelial lesion or malignancy" were compared with cases of "true ASCUS," using histologic squamous intraepithelial lesion and/or a positive high-risk HPV test as a positive outcome (abnormal follow-up).

Of 100 cases, 28 were reclassified as benign by both observers. In 15 of these, the original ASCUS interpretation was based on cells with bland nuclear enlargement (2-3 times the area of intermediate cell nuclei), smooth nuclear membranes, and fine chromatin. Abnormal follow-up was identified in 1 (7%) of 15 benign cases but in 30 (42%) of 72 true ASCUS cases (P = .023). PM cells are a significant cause of ASC overdiagnosis in women 40 to 55 years old.

Cervical Paps with cells no more atypical than these can be interpreted safely as negative for intraepithelial lesion or malignancy.

The Small Blue Cell Dilemma Associated With Tamoxifen Therapy

Yi Jun Yang, MD, PhD, Linda K. Trapkin, DO, Roberta K. Demoski, CT(ASCP), Jeannette Bellerdine, MS, SCT(ASCP), and Celeste N. Powers, MD, PhD

From the Department of Pathology, Upstate University, State University of New York, Syracuse (Dr Yang and Ms Demoski); the Department of Pathology, Laboratory Alliance of Central New York at St Joseph's Hospital, Syracuse (Dr Trapkin and Ms Bellerdine); and the Department of Pathology, Medical College of Virginia, Virginia Commonwealth University, Richmond (Dr Powers)

Arch Pathol Lab Med 2001;125:1047–1050. Abstract quote

Context. —Several endometrial diseases, such as endometrial hyperplasia, endometrial carcinoma, and endometrial polyps, have been reported to be associated with tamoxifen administration. We recently observed a high incidence of distinctive small blue cells in Papanicolaou tests of women who had received tamoxifen treatment for breast carcinoma.

Objectives. —To define the characteristics of these small blue cells, to identify the patient population in which they are found, and to determine the clinical significance and possible etiology of these findings.

Design.—A total of 154 Papanicolaou tests from 60 patients with a clinical history of tamoxifen therapy were reviewed retrospectively.

Results. —Small blue cells were found in 40% of Papanicolaou tests from patients who received tamoxifen therapy. Patients with small blue cells in their Papanicolaou tests were an average of 9 years older at the time tamoxifen therapy was initiated than those without. Among the available follow-up surgical biopsies, no malignant diagnoses were made.

Conclusions. —We conclude that these distinctive small blue cells are found more frequently in older patients and most probably represent proliferative reserve cells of cervical/vaginal epithelium resulting from the estrogenic agonist effect of tamoxifen. More importantly, they are nonneoplastic in nature.


Misdiagnosis of High-Grade Vulvar Intraepithelial Neoplasia (VIN III) as Mild Cervical Intraepithelial Neoplasia (CIN I) on Papanicolaou Tests.

August CZ, Ganji M, Froula E.

Department of Pathology, Advocate Illinois Masonic Medical Center, Chicago, Ill.

Arch Pathol Lab Med 2003 Jan;127(1):68-70 Abstract quote

Context.-Although high-grade vulvar intraepithelial neoplasia (VIN III) is a clinically significant lesion, it can be overlooked because of nonspecific clinical findings and the fact that its cytomorphologic features mimic those of mild cervical intraepithelial neoplasia (CIN I).

Objective.-To determine if there are cytomorphologic features on Papanicolaou tests that can reliably distinguish between VIN III and CIN I.

Design.-Papanicolaou tests diagnosed as CIN I from patients with biopsy-proven CIN I were compared with Papanicolaou tests diagnosed as CIN I from patients with biopsy-proven VIN III but with no biopsy-proven CIN I.

Results.-None of the cytomorphologic features evaluated could reliably distinguish CIN I from VIN III.

Conclusion.-Since the Papanicolaou test cannot be used to distinguish between CIN I and VIN III, the clinician must pay careful attention to the clinical and colposcopic findings. Further research evaluating the use of ancillary studies, such as human papillomavirus typing, may be useful.



Significance of AGUS Pap smears in pregnant and postpartum women.

Chhieng DC, Elgert P, Cangiarella JF, Cohen JM.

Department of Pathology, New York University Medical Center, New York, New York, USA.

Acta Cytol 2001 May-Jun;45(3):294-9 Abstract quote

OBJECTIVE: To study the clinical significance of atypical glandular cells of undertermined significance (AGUS) in pregnant and postpartum women.

STUDY DESIGN: We evaluated 35 women who were pregnant (30) or within three months postpartum (5) and had a cytologic diagnosis of AGUS. Twenty-seven (77%) patients had follow-up: 17 (63%) patients underwent colposcopic examination and biopsy, and 10 (37%) had repeat Pap smears. Eight patients were lost to follow-up.

RESULTS: Five (29.4%) patients had a squamous intraepithelial lesion (SIL), including three high grade and two low grade, on subsequent biopsy. The remaining (70.6%) patients had benign pathology, which included 5 chronic cervicitis, 4 endocervical and/or endometrial polyps, 2 Arias-Stella reaction and 1 microglandular hyperplasia. Among the patients with repeat Pap smears, two had persistent AGUS/atypical squamous cells of undetermined significance, the remaining cases were within normal limits.

CONCLUSION: Pregnancy-related changes may present with glandular atypia. In addition, about one-third of pregnant and postpartum women with a diagnosis of AGUS had SIL on subsequent biopsy; that rate is similar to that in nonpregnant women. Therefore, pregnant women with a cytologic diagnosis of AGUS should be followed closely.

Usefulness of DNA Ploidy Measurement on Liquid-Based Smears Showing Conflicting Results Between Cytology and High-Risk Human Papillomavirus Typing

Marianne Lorenzato, PhD,1 Jean-Paul Bory, MD,2 Joël Cucherousset, MD,1 Jean-Marie Nou, MD,1 Dominique Bouttens,1 Corinne Thil,1 Francine Dez,1 Ghislaine Evrard,1 Christian Quereux, MD,2 Philippe Birembaut, MD,1 and Christine Clavel, PhD

Am J Clin Pathol 2002;118:708-713 Abstract quote

To improve the positive predictive value (PPV) for high-risk human papillomavirus (HR-HPV) in primary screening, DNA ploidy was measured on the same liquid-based sample by image cytometry in 984 cases showing discrepancies between cytology and HR-HPV testing.

Of the conflicting results, 14.5% corresponded to a cytologic lesion (from atypical squamous cells of undetermined significance to high-grade squamous intraepithelial lesion [HSIL]) without HPV detected, and 85.5% of smears were within normal limits but revealed an HR-HPV infection. A suspect DNA profile was associated significantly with a lesion. In 497 patients who underwent repeated HPV testing, a normal DNA profile at the first smear predicted the clearance of HPV infection (sensitivity, 81.5%; specificity, 45.4%; PPV, 69%; negative predictive value, 62.4%).

In persistent HR-HPV infection, a suspect DNA profile at the first smear increased the PPV from 10.8% to 22.7% for the detection of a histologically proven HSIL with a sensitivity of 95.2%. DNA ploidy can be used to select smears with high risk of HSIL, especially in cases of persistent HR-HPV infection.


Improving Pap test turnaround time using external benchmark data and engineering process improvement tools.

Persoon TJ, Zaleski MS, Cohen MB.

Department of Pathology, University of Iowa, Iowa City 52242, USA.


Am J Clin Pathol 2002 Oct;118(4):527-33 Abstract quote

Turnaround time for Papanicolaou (Pap) tests became an important service quality issue at our institution.

We studied Pap test turnaround time using engineering process improvement tools and benchmarked turnaround time against data published as a College of American Pathologists Q-Probes study. An IDEF3 process map revealed the complexity of the Pap test process and the opportunities for process improvement.

We used these data and the action-research method to initiate changes in cytopathology laboratory operations with the goal of reducing turnaround time. Before intervention, mean Pap test turnaround time was highly variable; during a 6-month period, monthly means ranged from 2.5 to 10.8 days. A cycle time study conducted over a 2-week period validated these data. After system improvements were implemented, the monthly mean turnaround time decreased and became more consistent, with 11 of 12 months having a mean turnaround time of 3 days or less (range, 1.5-3.9 days).

Our study illustrates the value of publishing Q-Probes data for use as external benchmarks and the benefits of using tools from other disciplines to improve laboratory processes.

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Commonly Used Terms with the Pap Smear

AGUS-Atypical Glandular Cells of Undetermined Significance.

ASCUS-Atypical Squamous Cells of Undetermined Significance.

Bethesda Classification 2001-This is the current diagnostic classification system for Pap smears.  It was created by a multidisciplinary committee to bring uniformity to the reports.  The most recent recommendation are found here.

Cervical Dysplasia and Cancer

CIN-This is cervical intraepithelial neoplasia.  It is a grading system for squamous dysplasia.  CIN I is termed mild dysplasia while CIN III is termed high grade dysplasia.  These terms have largely been replaced by the Bethesda Classification.


Human Papilloma Virus-The etiologic agent for condyloma and cervical cancer.

Koilocyte-This is a common cytologic change seen in the cervical squamous epithelial cells after infected with human papilloma virus.  It is sometimes described as raisinoid after its wrinkled hyperchromatic appearance.

Transformation Zone-This is the area in the cervix where most cases of dysplasia and cancers arise. 

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Last Updated September 12, 2007

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