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The normal adult red blood cell lacks a nucleus; it is gradually lost just before its release from the bone marrow.  In disease states, nucleated red blood cells may be found in the circulation. The peripheral smear is made by placing a drop of blood on a slide and smearing it to a fine layer. It is then stained with a Wright-Giemsa stain and examined under the microscope. A sample of blood can also be placed in an automated analyzer which sorts the cells by size and amount of light scatter. The result is a complete blood count (CBC) which also gives the quantities of the white blood cells and platelets.


Alpha Thalassemia
Beta Thalassemia
Paroxysmal Nocturnal Hemoglobinuria (PNH)
Aplastic Anemia
Sickle Cell Anemia

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ESR (Erythrocyte Sedimentation Rate)


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Cross-Reactivity of Cefotetan and Ceftriaxone Antibodies, Associated With Hemolytic Anemia, With Other Cephalosporins and Penicillin

Patricia A. Arndt, MS, MT(ASCP)SBB, and George Garratty, PhD, FRCPath

Am J Clin Pathol 2002;118:256-262 Abstract quote

Most drug-induced immune hemolytic anemias since the late 1980s have been caused by the second- and third-generation cephalosporins, cefotetan and ceftriaxone, respectively. Cross-reactivity of cefotetan and ceftriaxone antibodies with other cephalosporins or penicillin has been studied only minimally.

We tested 7 serum samples previously identified to contain cefotetan antibodies and one serum sample previously identified to contain ceftriaxone antibodies against 9 other cephalosporins, penicillin, and 7-amino-cephalosporanic acid in the presence of RBCs and also used hapten inhibition to indicate cross-reactivity. Serum samples containing cefotetan antibodies showed some cross-reactivity with cephalothin and cefoxitin (and to a much lesser extent with penicillin and ceftazidime). The ceftriaxone antibodies showed very weak cross-reactivity with cefotaxime, cefamandole, and cefoperazone. There was very little cross-reactivity between cefotetan antibodies and the drugs tested in the present study.

We have no data to determine whether the in vitro data relate to in vivo reactivity.


Flow Cytometric Measurement of Hemoglobin F in RBCs
Diagnostic Usefulness in the Distinction of Hereditary Persistence of Fetal Hemoglobin (HPFH) and Hemoglobin S–HPFH From Other Conditions With Elevated Levels of Hemoglobin F

James D. Hoyer, MD, Connie S. Penz, Virgil F. Fairbanks, MD, Curtis A. Hanson, MD, and Jerry A. Katzmann, MD


Am J Clin Pathol 2002;117:871-879 Abstract quote

The cellular distribution of hemoglobin F is important for evaluating persistently elevated hemoglobin F levels, such as in hereditary persistence of fetal hemoglobin (HPFH) or delta/beta-thalassemia, and for differentiating homozygous hemoglobin S (or hemoglobin S–beta0-thalassemia) from hemoglobin S–HPFH, traditionally done by using the Kleihauer-Betke (K-B) acid elution test.

We evaluated a flow cytometric method using an anti–hemoglobin F antibody as a replacement for the K-B test. We used 172 specimens representing a variety of conditions: HPFH trait, 19 cases; delta/beta-thalassemia trait, 8 cases; hemoglobin S–HPFH, 10 cases. By flow cytometry, all cases of HPFH trait gave a hemoglobin F pattern comparable to the homocellular pattern obtained by the K-B test; all cases of delta/beta-thalassemia tested gave a pattern comparable to a K-B heterocellular pattern. Most cases of hemoglobin S–HPFH gave a homocellular distribution of hemoglobin F, whereas all cases of homozygous hemoglobin S with elevated hemoglobin F levels gave a heterocellular pattern.

Flow cytometry provides a more rapid and objective method for assessing cellular distribution of hemoglobin F and is useful for patient evaluation when HPFH trait, delta/beta-thalassemia trait, or hemoglobin S–HPFH trait is suspected.

Noninvasive Monitoring of Hemoglobin The Effects of WBC Counts on Measurement

Katsuyasu Saigo, MD, Shion Imoto, MD, Makoto Hashimoto, Hisashi Mito, MD, Junko Moriya, MD, Tadanobu Chinzei, MD, Yoshitsugu Kubota, MD, Shigehiro Numada, Toshiyuki Ozawa, Shunichi Kumagai, MD
Am J Clin Pathol 2004;121:51-55 Abstract quote

The efficacy of a noninvasive hemoglobin monitoring device (Astrim, Sysmex , Kobe, Japan) was evaluated for healthy volunteers and for patients with hematologic disorders. At the same time, the effects of WBC counts on noninvasive monitoring were studied by clinical evaluation and in ex vivo experiments.

The hemoglobin levels determined by the device (Ast-Hb) and a conventional analyzer (T-Hb) were compared. The coefficient of correlation between findings with the Ast-Hb and the T-Hb for healthy volunteers was r = 0.626, whereas that for patients with hematologic disorders was r = 0.762. A comparison of the ratios of measurement errors in hemoglobin levels by Ast-Hb and T-Hb indicated that the number of WBCs had no effect on hemoglobin monitoring.

Moreover, ex vivo studies using isolated WBCs and an optical model that imitates blood vessels and tissue in human fingers confirmed these results. Therefore, this new hemoglobin monitoring device can be expected to be useful for continuous hemoglobin monitoring.
Standardization of Reticulocyte Values in an Antidoping Context

Michael J. Ashenden, PhD, etal.
Am J Clin Pathol 2004;121:816-825 Abstract quote

The lack of standardization of reticulocyte results hinders the ability of sports authorities to recognize the telltale fluctuations over time that are typical for athletes using illegal blood doping to improve their performance. Therefore, the aim of the present study was to devise a tenable approach for antidoping authorities to quantify instrument bias.

We evaluated reticulocyte data derived during a 42-week period from 210 hospital patient blood samples measured in duplicate simultaneously on up to 11 hematology analyzers located in a single laboratory.
We found that square root transformation of reticulocyte values enabled quantification of interinstrument bias by using the mean reticulocyte value of a cohort of approximately 54 subjects as a de facto calibration agent.

We also demonstrated that measurement precision associated with low reticulocyte values was not inferior to that associated with higher values.
The New Reticulocyte Parameter (RET-Y) of the Sysmex XE 2100
Its Use in the Diagnosis and Monitoring of Posttreatment Sideropenic Anemia

Mauro Buttarello, MD, etal.
Am J Clin Pathol 2004;121:489-495 Abstract quote

To verify their clinical usefulness in diagnosis and early response to therapy of sideropenic anemia, we compared the behavior of the reticulocyte parameter (RET-Y), a raw measure dependent on size and content of the cell, generated by the Sysmex XE 2100, with the mean reticulocyte volume (MCVr) and mean reticulocyte hemoglobin content (CHr) from the Bayer ADVIA 120 in healthy subjects and patients with iron deficiency anemia. Correlations were high (r = 0.88 and r = 0.94, respectively). All parameters varied significantly as early as 48 hours after the start of intravenous iron therapy (mean differences of 17.4% [RET-Y], 4.5% [MCVr], and 9.5% [CHr]). Sudden decreases in those parameters at interruption of therapy indicate the reappearance of sideropenic erythropoiesis.

The receiver operating characteristic curve demonstrated a high degree of efficiency in differentiating moderate or severe iron deficiency anemia from the healthy state. The best association between sensitivity and specificity was at a cutoff of channel 1,624 for RET-Y and 104.5 fL for MCVr (negative and positive predictive values, respectively, of 99.6% and 96.5% for RET-Y and 98.7% and 93.3% for MCVr).

RET-Y is correlated closely with CHr and is useful for diagnosis and early monitoring after the administration of intravenous iron.
Five Fully Automated Methods for Performing Immature Reticulocyte Fraction
Comparison in Diagnosis of Bone Marrow Aplasia

Mauro Buttarello, MD,1 Pietro Bulian, MD,1 Giorgio Farina, MD,1 Maria Grazia Petris, MD,2 Valeria Temporin, MD,1 and Lucia Toffolo, MD

Am J Clin Pathol 2002;117:871-879 Abstract quote

We performed a parallel evaluation of 5 automated reticulocyte counters to produce the immature reticulocyte fraction (IRF).

We analyzed 225 samples from healthy control subjects, 115 from patients with various diseases, 38 with advanced aplasia, and 22 in early erythropoietic recovery after chemotherapy or bone marrow transplantation. The reference intervals were different for each instrument (ADVIA 120, 0.04-0.25; CELL DYN 4000, 0.15-0.35; GEN-S, 0.20-0.37; SE 9500 RET, 0.05-0.21; VEGA RETIC: 0.06-0.23). The imprecision, obtained by 1-way analysis of variance on duplicates, was satisfactory for clinical use for all methods (coefficient of variation, 7.6%-20.5% in healthy subjects), although it was higher than the analytic goal based on biologic variability within subjects.

The comparison of different methods shows that agreement is good only between SE 9500 RET, CELL DYN 4000, and VEGA RETIC (r2 = 0.72-0.78). The study of diagnostic performance in distinguishing aplasia from early bone marrow recovery shows slightly different results (area under the curve from 0.70 for ADVIA 120 to 0.96 for SE 9500 RET). Even with slight differences, the fluorescence-based methods seem to be more robust than other methods for IRF measurement.

Henry JB. Clinical Diagnosis and Management by Laboratory Methods. Twentieth Edition. WB Saunders. 2001.

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Last Updated 6/11/2004

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