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One tube with eight antibodies for 14‐part bone marrow leukocyte differential using flow cytometry
Author(s) -
Jacob MarieChristine,
Souvignet Alice,
Pont Julie,
Solly Françoise,
Mondet Julie,
Kesr Sanae,
Pernollet Martine,
DumestrePerard Chantal,
Campos Lydia,
Cesbron JeanYves
Publication year - 2017
Publication title -
cytometry part b: clinical cytometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.646
H-Index - 61
eISSN - 1552-4957
pISSN - 1552-4949
DOI - 10.1002/cyto.b.21369
Subject(s) - flow cytometry , bone marrow , pathology , cytometry , cd34 , cd19 , antibody , immunophenotyping , immunology , biology , medicine , stem cell , genetics
Background Bone marrow analysis by flow cytometry is part of the routine diagnosis of hematological disorders in medical laboratories. Differential leukocyte count and identification of abnormal cell subsets is currently performed through morphological examination on bone marrow smears by skilled cytologists. In this work, we propose a single 8‐color tube for providing equivalent information, using flow cytometry. Methods 99 bone marrow samples were classified into 2 groups, (i) 51 samples, obtained from either healthy donors ( n = 4) or patients with various diseases at diagnosis or during remission that did not present a hematological malignancy ( n = 47), and (ii) 48 pathological samples with quantitative and/or qualitative abnormalities. A panel of eight antibodies—CD3‐FITC/CD10‐PE/CD38‐PerCP‐Cy5.5/CD19‐PECy7/CD36‐APC/CD16‐APC‐H7/CD34‐BV421/CD45‐V500—was tested to identify the main cell subsets at different stages of maturation using a FACSCanto‐II analyzer. Results We first proposed a strategy of sequential gating leading to the identification of 14 leukocyte subsets, that is, erythroblasts, monocytes, B‐lymphoid cells from hematogones to plasma‐cells (5 subsets), T‐ and NK‐cells, polymorphonuclear cells (neutrophils, eosinophils, and basophils), myeloblasts and other immature granular cells. This approach was validated by comparing flow cytometry and microscopic morphological examination, both in cases of normal and abnormal samples. Interestingly, cell identification, and numeration by flow cytometry was easy to perform and highly reproducible. Conclusion A very simple, rapid, and reproducible flow cytometric approach, using a combination of eight antibodies allows determination of the cellular composition of bone marrow with high precision. © 2016 International Clinical Cytometry Society