Antibodies directed against CD20 (L26, Leu 16, and Bl) are frequently used to determine the presence of B lymphocytes. However, recent publications describe the unexpected presence of CD20-positive T cells in the peripheral blood of normal subjects and occasional T-cell neoplasms that express CD20. To determine the presence of CD20-positive T cells in bone marrow, flow cytometric analysis was performed on 34 aspirate specimens (14 normal, 5 acute lymphoblastic lymphoma |ALL|, 5 acute myelogenous leukemia |AML|, 4 HIV positive, 2 myelodysplastic/myeloproliferative, 2 chronic myelogenous leukemia |CML|, 1 chronic lymphocytic lymphoma |CLL], 1 multiple myeloma). A small population of cells coexpressing CD3 (Leu 4) and CD20 dim (Leu 16) was identified in 94% of the specimens, representing 0% to 11%Phenotyping of human lymphoid proliferations is of value in determining cell lineage. Immunophenotyping uses monoclonal antibodies directed against B-or T-cell specific antigens. B lymphocytes are identified by the presence of the antigens CD 19, CD20, CD21, and CD22. CD20 is a 33-kDa membrane phosphoprotein' with three hydrophobic regions that traverse the cell membrane, creating a structure similar to an ion channel. 2This protein is expressed on the majority of B cells, appearing early in B-cell development (following CD 19 and CD 10, but preceding CD21, cell surface expression of CD22, and surface immunoglobulin). "5 CD20 is retained on mature B cells until plasma cell development. 34 The CD20 antigen can be identified by immunohistochemistry or flow cytometry using several commercially available monoclonal antibodies: Leu 16, Bl, and L26 (Leu 16

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Flow cytometric analysis is important in the diagnosis, classification, and follow-up of non-Hodgkin lymphoma. It is assumed that the lymphoma phenotype for each patient remains unchanged over time and is consistent from one specimen to another. To determine the variability in expression of lymphoid antigens, we reviewed 211 flow cytometry specimens of malignant lymphoma from 81 patients. Some antigens showed a stable pattern of expression such as CD5, CD10, CD19, CD20, and HLA-DR. In contrast, CD21, CD22, CD23, and CD25 showed more variability from one specimen to another. We believe several factors affect the stability of antigen expression. True differences in expression most probably are related to the biology and function of the different antigens. For instance, CD19 and CD20 are essential in cell maturation and function and, therefore, are present on the majority of cells. In contrast, CD22 has a role during B-cell activation and, therefore, is more variable. Lack of standardization inflow cytometry procedures also is responsible for some variability. Instrument settings for adequate compensation and the criteria used to determine when an antigen is reported as positive are important considerations when evaluating flow cytometry histograms.