Flow cytometric detection of minimal residual CLL Detection using the protocol published by Rawstron and colleagues.¹⁰  Top row of plots demonstrate preliminary gating on CD19⁺ B cells with low orthogonal (side) light scatter and refinement of the gate using a plot of forward versus low orthogonal (side) light scatter. Bottom 3 rows of plots demonstrate representative dot-plots from 3 separate tubes evaluating CD5, CD20, and CD38, (second row); CD5, CD22, and CD81 (third row); and CD5, CD43, and CD79b (bottom row). Only the CD19⁺ gated events are displayed. Regions are placed around events with phenotypic features characteristic of CLL: R5 indicates dim CD20 and CD5⁺; R6, dim CD20 and dim to moderate intensity CD38 staining; R7, CD5⁺ and CD22⁺; R8, CD22⁺ and dim CD81; R9, CD5⁺; and CD79b^dim, R10, CD5⁺ and CD43⁺. Residual CLL is identified by more than 50 events meeting the criteria for 2 or more of the following: R5 and R6 (displayed in second row right dot plot), R7 and R8 (displayed in third row right dot plot), R9 and R10 (displayed in bottom row right dot plot). The flow cytometric data were acquired using a BD FACS Calibur flow cytometer (BD Biosciences) and the dot plots were created using BD CellQuest software v3.3 (BD Biosciences).