Kolmogorov-Smirnov analysis for identification of antibodies that differentiate between normal and neoplastic B cells.

(A) Kolmogorov-Smirnov analysis was used to identify the degree of separation provided by the panel of antigens assessed. Briefly, the cumulative fluorescence intensity of a particular antigen is plotted for both normal B cells and CLL cells. The D value is the maximum difference between these 2 distributions (1 = complete separation, 0 = complete overlap), and is used as a simple measure of the degree of difference in expression. The plot demonstrates the low level of overlap between normal and neoplastic CD20 expression, in spite of the fact that both antigens are “positive” compared with isotype control. (B) Antibodies for optimal separation of CLL cells from normal B cells. The figure shows the degree of separation (K-S D value) for a panel of antibodies assessed on normal B cells and on presentation CLL cells. Values shown are the 5th, 25th, median, 75th, and 95th percentiles, and antibodies are ranked according to median D value (worst separation on the left, best separation on the right). Antigens that are expressed at a higher level by CLL cells than by normal B cells have an asterisk above (CD5 and CD23), whereas those expressed at a lower level by CLL cells are not marked. Peripheral blood antigen expression is shown in (i)—CD10 is expressed by neither normal peripheral blood B cells nor by CLL cells, and this demonstrates the differences due to autofluorescence. Bone marrow antigen expression is shown in (ii).