Bcl-2 expression and its prognostic value were evaluated in 42 children with acute leukemia. The Bcl-2 expression of the leukemic blast cells was measured quantitatively by flow cytometry and was further analyzed by the simultaneous immunostaining of Bcl-2 with the surface membrane antigens, DNA, Ki-67 antigen. All of the cases showed a consistent expression of Bcl-2 protein; virtually all leukemic lymphoblasts were Bcl-2 positive. Although the expression of Bcl-2 varied widely from 7 to 80 x 10(3) MESF units, no significant difference was found in the mean value between the patients with acute lymphoblastic leukemia and those with acute myeloblastic leukemia. In more than half of the patients with AML, intraclonal heterogeneity of Bcl-2 expression was observed. The expression of Bcl-2 showed no apparent fluctuations during the different phases of the cell cycle. However, the proportion of Bcl-2-positive and -negative cells during the cell cycle was different between ALL and AML patients. In the ALL patients, few Bcl-2-negative cells were detected only in the GI phase, whereas in the AML patients Bcl-2-negative cells were detected in the S and G2/M phases, as well as in the G1 phase. No apparent difference was found in Bcl-2 expression between the Ki-67-negative noncycling population and the Ki-67-positive cycling population. Of the clinical features of these patients, only CD34 expression in the ALL patients was associated with high levels of Bcl-2 expression. In the 28 untreated cases of ALL, high expression of Bcl-2 was not an unfavorable factor for the outcome of this disease.