Colony-forming assay for circulating chronic lymphocytic leukemia cells.

In these studies, we report adaptation of a colony-forming assay to chronic lymphocytic leukemia (CLL) peripheral blood cells. T-lymphocyte-depleted CLL peripheral blood cells were cultured with irradiated, normal T cells and media conditioned by normal, mitogen-stimulated T cells in methylcellulose. Colonies containing small and transformed lymphocytes appeared after 5-7 days incubation. The plating efficiency of CLL colonies was 0.15 +/- 0.08% (x +/- S.D.), similar to that of other colony-forming systems. The majority of CLL colony-forming cells were in S phase (50 +/- 4% x +/- S.E.) as determined by thymidine suicide and the fraction of colony-forming cells in S phase was inversely related to the WBC. Cells harvested from CLL colonies lacked surface markers for T lymphocytes and stained positively for monoclonal surface and/or cytoplasmic immunoglobulin light chains. A 1-h incubation was used to study the in vitro response of CLL colony-forming cells to adriamycin and melphalan. Preliminary studies suggest differences in patterns of in vitro sensitivity to melphalan between patients previously treated with alkylating agents and those who had not received treatment. This system can be used to study regulation of CLL cell proliferation, and may have utility in predicting response to chemotherapeutic agents.