Reticulocyte analysis using light microscopy and two different flow cytometric procedures.
The reticulocyte count is a clinically important indirect indicator of erythropoietic activity of the bone marrow. Reticulocyte enumeration by light microscopy is rather inaccurate and has poor reproducibility. Automation of the reticulocyte count by means of flow cytometry has considerably improved the quality of this investigation. In our study, we compared three methods of establishing the blood reticulocyte number: the microscopic brilliant cresyl blue method and two flow cytometric procedures using thiazole orange (TO), namely FACSort (Becton-Dickinson) and EPICS Profile (Coulter). The aims of the study were (1) to select the most suitable TO concentration to be used with the EPICS Profile cytometer, (2) to determine the correlation between the microscopic method and the two flow cytometric procedures, and (3) to appraise the suitability of flow cytometry for reticulocyte analysis in routine clinical work. According to our results, the most appropriate TO concentration for the EPICS Profile counter is 0.1 mg/L. We observed a good correlation between the three methods tested; the correlation coefficients ranged from 0.82 to 0.87. The mean intra-assay coefficients of variation for the microscopic method and the EPICS Profile and FACSort procedures were 27.5%, 8.4% and 6.3%, respectively.
Preloznik-Zupan
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Department of Haematology, University Medical Centre, Ljubljana, Slovenia.
Cernelc
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Zontar
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D052160:Benzothiazoles; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes / Q000008:administration & dosage; D006801:Humans; D009994:Osmolar Concentration; D011804:Quinolines; D012016:Reference Values; D017701:Reticulocyte Count / Q000379:methods*; D012156:Reticulocytes / Q000166:cytology*; D013844:Thiazoles / Q000008:administration & dosage; D013997:Time Factors
D052160:Benzothiazoles; D005456:Fluorescent Dyes; D011804:Quinolines; D013844:Thiazoles; C051000:thiazole orange