Flow cytometric analysis of BTK expression in blood leukocytes from normal donors. (A) Two-color immunofluorescence analysis of BTK expression in B cells and non-B cells. Mononuclear cells were stained with PE-labeled anti-CD20 MoAb, fixed, and permeabilized. The cells were then reacted with anti-BTK or irrelevant control MoAbs and further incubated with FITC-labeled secondary antibody. The dot plot map (a) shows the two-color pattern of lymphocytes gated by forward and right angle right scatter. A second gate was set to include CD20⁺ B cells (b) and non-B cells (c), and the BTK expression in each cell population is presented as a histogram. (B) Analysis of BTK expression in monocytes and neutrophils. Mononuclear cells or neutrophils were stained for BTK as described above. BTK expression in monocytes was evaluated by gating on CD14⁺ population. The dashed line indicates the control antibody. Five thousand cells were evaluated in each gated population.