Flow cytometric differential of leukocyte populations in normal bone marrow: Influence of peripheral blood contamination

Background: Availability of immunophenotypic reference values for the various leukocyte populations distributed in bone marrow may be helpful to recognize abnormal bone marrow development and, therefore, useful as first screening of individuals with suspected hematological malignancies or other hematopoietic disorders. Methods: A single tube four-color staining panel (CD66abce/CD14/CD45/ CD34) together with a predefined gating strategy was utilized to immunologically differentiate the distribution of the major leukocyte populations in bone marrow aspirates of healthy donors. The sample-blood erythrocyte ratio was applied to assess the amount of blood contamination of marrow and account for this in the marrow value estimates. Results: The frequency of the major leukocyte populations in bone marrow of 134 normal donors were for granulocytes: mean, 69.4%; SD, 10.3%; monocytes: mean, 4.7%; SD, 2.3%; lymphocytes: mean, 18.3%; SD, 8.7%. The frequency of the immature cell population that included precursor cells of each of the cell lineages among other cell types were mean 5.0%; SD 2.2%. The mean percentage of CD34 positive cells was 1.5%; SD 0.7%. Our results showed further that the frequency of cell populations, of which the presence is restricted to the bone marrow (e.g., CD34 + progenitor cells), is influenced by the degree of peripheral blood admixture. Between the total immature cells and purity of the bone marrow, there was a significant positive correlation demonstrated, whereas a negative correlation was found between the percentages of both lymphocytes as monocytes and the purity of the bone marrow. Conclusions: With a single tube-staining panel, we obtained reference values for flow cytometric assessment of all relevant leukocyte populations present in bone marrow that can be used as a frame of reference for better recognition of individuals with abnormal hematopoiesis. In addition, we have demonstrated the influence of the degree of peripheral blood admixture in the bone marrow aspirates on those reference values.

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