Flow cytometric immunophenotypic analysis of systemic mastocytosis involving bone marrow

Context: Mast cells of systemic mastocytosis (SM) have aberrant immunophenotypes that are useful for their detection by flow cytometry immunophenotyping.

Objectives: To assess the usefulness of CD2, CD25, and other antigens for establishing the diagnosis of SM in bone marrow using flow cytometry immunophenotyping.

Design: We studied 50 bone marrow aspirates of patients with SM using flow cytometry immunophenotyping. The bone marrow aspirates were stained with antibodies specific for CD2, CD25, CD35, CD59, CD63, and CD69. For the detection of CD2 and CD25, antibodies conjugated with phycoerythrin (PE) or fluorescein isothiocyanate (FITC) were compared. CD45-PerCP and CD117-APC were used for gating. Data were acquired on FACS Calibur cytometers and analyzed using CellQuest software.

Results: CD2 and CD25 were positive in 41 of 50 (82%) and 45 of 50 (90%) SM cases, respectively. For CD2, the PE-conjugated antibody yielded better sensitivity than the FITC-conjugated antibody (31 of 40 [78%] versus 28 of 40 [70%]). For CD25, PE-conjugated and FITCconjugated antibodies showed similar detection sensitivity, although the intensity of expression was brighter with CD25-PE. Compared with immunohistochemistry, flow cytometry immunophenotyping was superior for detecting CD2 (14 of 23 [61%] versus 9 of 23 [39%]). Other antigens frequently overexpressed in SM were CD35 (43 of 50 [86%]), CD59 (46 of 50 [92%]), CD63 (43 of 49 [88%]), and CD69 (39 of 48 [81%]).

Conclusions: Flow cytometry immunophenotyping is a rapid and sensitive technique for characterizing mast cells in bone marrow aspirate specimens. The use of PEconjugated antibodies for CD2 and CD25 improves the detection rate (CD2) or facilitates analysis (CD25); therefore, PE-conjugated antibodies are suggested. Antibodies reactive with CD35, CD59, CD63, and CD69 are also helpful in detecting SM in bone marrow.