Cytoplasmic IgM in leukemic B cells by flow cytometry

The presence of intracellular cytoplasmic immunoglobulin M (IgM) in leukemic cells from patients with acute lymphocytic leukemia (ALL) and chronic lymphocytic leukemia (CLL) was investigated by flow cytometry. The objective of the study was to develop a reproducible flow cytometric method. A Burkitt's lymphoma-derived B-cell line, Daudi, and a pre-B ALL, Nalm-6, served as prototypes. Normal B cells and cells from patients with chronic myelogenous leukemia (CML) were used as negative controls. Cytoplasmic μ was expressed in 77.3 ± 7.5% (n = 10) of Nalm-6 cells. CALLA+ ALL and CML cells lacked cytoplasmic μ. The surface-membrane immunoglobulin on the viable B cells was blocked with purified goat anti-human IgM. Subsequently, the B cells were fixed in cold absolute methanol and stained with a fluorescein-conjugated goat anti-human IgM to demonstrate cytoplasmic IgM. After the surface-membrane IgM was blocked, normal B cells had no cytoplasmic IgM (0.3-0.5% positive cells) detectable by flow cytometry. However, the peripheral blood lymphocytes from five patients with CLL and the Daudi cells contained cytoplasmic IgM, ranging from 7.8 to 76.7% and 45.7 to 89.3%, respectively. We conclude that cytoplasmic μ in Nalm-6, CLL, and Daudi cells can be easily and rapidly demonstrated by flow cytometry.