FLOW-CYTOMETRY CD45 GATING FOR IMMUNOPHENOTYPING OF ACUTE MYELOID-LEUKEMIA

A flow cytometry method has been introduced into the routine investiga tion of whole bone marrow samples following red blood cell lysis on th e basis of a primary CD45/side scatter (SSC) gating procedure. Blast c ells were first identified by CD45/SSC gating in 74 cases of acute mye loid leukemia (AML) and the results were compared to a conventional FS C/SSC gating procedure and to MGG-staining smears. The percentages of blast cells in these samples as defined by the morphological analysis of MGG smears correlated better with the values determined by CD45/SSC gating (r = 0.94) than with the blast cell counts recorded with FSC/S SC gating (r = 0.76). These findings were not surprising because while CD45 expression was regularly lower on leukemic blasts than on normal lymphoid and monocytic cells, the FCS/SSC characteristics of these po pulations were overlapping. In 53 samples, the blast cell populations were also analyzed with a panel of FITC-conjugated monoclonal antibodi es that were utilized in double labeling with CD45-PE, We show that th e CD45/SSC gating procedure improved phenotypic determination of the b last cells in three ways: (1) by discriminating between leukemic blast cells and residual normal cells; (2) by excluding normal cells from t he phenotypic analysis of leukemic blast cells; and (3) by identifying blast cell heterogeneity in many cases of leukemia on the basis of di fferent CD45 display. Moreover, this immunophenotyping procedure on wh ole bone marrow samples also allowed an efficient discrimination betwe en the various cell lineages and facilitated the analysis of leukemic blasts present in low proportions.