In this study, the proliferative activity of human B and T cell precursors in central lymphoid organs, acute lymphoblastic leukemia (ALL) cells, and permanent cell lines was investigated with double- and triple-color-labeling methods for the analysis of cell cycle-associated features such as 5-bromo-2′-deoxyuridine (BrdU) incorporation and the expression of a nuclear proliferation-associated antigen, Ki67, together with the phenotypic profile of the cells. In infant and regenerating bone marrow (BM), 41.5% +/- 4.0% of terminal deoxynucleotidyl transferase (TdT+) cells were Ki67+, and 30.0% +/- 4.0% incorporated BrdU. A similar proportion of TdT+ dividing cells was observed in adult BM. The proliferative activity of the B cell progenitors reached the peak at the pre-B stage: 80.8% +/- 7.6% and 35.3% +/- 6.1% of c mu +, RFB7- cells were Ki67+ and BrdU+, respectively. In contrast, greater than 95% of surface immunoglobulin- positive BM lymphocytes were resting cells. In infant thymus the highest dividing capacity (95% Ki67+, 60% to 90% BrdU+) was observed in large cortical thymocytes (TdT+, CD1-, cCD3+), and TdT+, CD1+ cortical thymocytes also showed a high proliferative activity (74.3% +/- 2.3% Ki67+, 22.0% +/- 1.0% BrdU+), but TdT-, mCD3+ thymic lymphocytes were mainly resting cells (less than 5% Ki67+, less than 1% BrdU+). The proliferative activity of null and common ALL blasts was significantly lower than that of normal BM TdT+ cells (15.5% +/- 4.2% Ki67+, 6.2% +/- 2.1% BrdU+; P less than .001). Dividing ALL blasts were TdT+ and expressed surface antigens detected by CD10 and/or CD19 antibodies. In T cell-ALL, the percentages of Ki67+ and BrdU+ blasts were also lower than those found in the corresponding normal immature thymocytes (13.0% +/- 3.1% and 2.4% +/- 1.3%, respectively; P less than .001). Thus, ALLs derive from actively proliferating lymphoid precursors but have a lower dividing capacity than the corresponding normal cell types. In ALL cases with heterogeneous expression of markers such as cmu and CD1, dividing blasts were distributed among both negative and positive populations, thus indicating that blasts with signs of differentiation also remain within the dividing pool.