Toward a new reference method for the leukocyte five-part differential
| Autor: | Johannes Streicher, Sylvia Andert, Peter Michael Bayer, Alexander Lapin, Wolfgang Hübl, Christin Hübner, Georg Wolfbauer, Gabriele Thum |
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| Rok vydání: | 1997 |
| Předmět: |
Biophysics
Biology Pathology and Forensic Medicine Flow cytometry Leukocyte Count Endocrinology Monocyte count Antigens CD Reference Values medicine Leukocytes Humans Sample preparation Chromatography medicine.diagnostic_test Antibodies Monoclonal Reproducibility of Results Liter Cell Biology Hematology Flow Cytometry Cell loss Flow (mathematics) Absolute count Fluorescent Antibody Technique Direct Reference values Immunology Regression Analysis |
| Zdroj: | Cytometry. 30(2) |
| ISSN: | 0196-4763 |
| Popis: | A flow cytometric method performing a five-part leukocyte differential based on three-color staining with anti-CD45-fluorescein isothiocyanate (FITC), anti-CD-14-phycoerythrin (PE)/Cy5, and a cocktail of PE-labeled anti-CD2, anti-CD16, and anti-HLA-DR antibodies was evaluated. Results obtained by using three different sample preparation procedures and two different flow cytometers were compared with those of a 1,000-cell manual differential for evaluation of accuracy. We observed excellent correlations with the manual differential for all leukocyte subclasses and even higher correlations between the different flow cytometric methods. Flow cytometric basophil results were identical to the manual counts, regardless of which sample preparation technique or flow cytometer was used. Therefore, we propose our flow cytometric method as the first acceptable automated reference method for basophil counting. The flow cytometric results for the other leukocyte subclasses were apparently influenced by the sample preparation, which could not be explained by cell loss during washing steps. Moreover, a small influence of the flow cytometer was also observed. Assessing the influence of sample storage, we found only minimal changes within 24 h. In establishing reference values, high precision of flow cytometric results facilitated detection of a significantly higher monocyte count for males (relative count: 7.08 +/- 1.73% vs. 6.44 +/- 1.33%, P < 0.05; absolute count: 0.536 +/- 0.181 x 10(9)/liter vs. 0.456 +/- 139 x 10(9)/liter, P < 0.01). Our data indicate that monoclonal antibody-based flow cytometry is a highly suitable reference method for the five-part differential: It also shows, however, that studies will have to put more emphasis on methodological issues to define a method that shows a high interlaboratory reproducibility. |
| Databáze: | OpenAIRE |
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