Proliferation and differentiation of normal and leukemic lymphocytes as analyzed by flow cytometry

Three different flow cytometric techniques are described and their application in studies of human lymphocyte proliferation and differentiation is reviewed. The first technique, simultaneous staining of DNA and RNA in addition to analysis of cell cycle distribution, reveals differences in the content of RNA in various types of normal, fully differentiated lymphocytes (B, Tμ, Tγ 'Null' cells). Lymphocyte dedifferentiation (blastogenesis, stimulation) is accompanied by an increase in RNA content, which is of higher degree in the case of B-cell than T-cell stimulation. G₀/G₁ lymphocytes from leukemia patients show differences in the content of RNA depending on the type of leukemia; RNA and DNA content has been found to be an useful parameter in classification of leukemias. In the second technique, incorporation of 5-bromodeoxyuridine (BUdR) into DNA by cycling lymphocytes can be detected and thus may be either used to assay cell proliferation or to label the mitogen responsive subpopulations ('memory cells') for further studies of secondary stimulation. In the third technique, the differences in the degree of nuclear chromatin condensation are used to discriminate fully differentiated, G₀ cells from stimulated lymphocytes, as well as to analyze mitotic activity in proliferating cell populations.