The effect of 1-beta-D-arabinofuranosylcytosine (ara-C) on mouse L1210 ascites tumor cells was studied in vivo with cell kinetic methods. Injection of ara-C (200 mg/kg) leads to an immediate inhibition of DNA synthesis and, thus, to a block of G1 cells at the G1-S border. This dose of ara-C kills a considerable fraction (at least 54%) of the L1210 cells. A double-labeling experiment with [3H]thymidine and [14C]thymidine which permits the distinction of cells in different cycle phases shows that ara-C kills the majority of cells in S phase, while only a minor fraction of G1 cells are lethally damaged. A multicompartment model is utilized to derive from the results of the double-labeling experiment the rates of cell loss out of the different cycle phases. Eighty-seven percent of the cells that are in S phase at the time of ara-C application and 21% of the cells in G1 are killed by ara-C. The repopulation of the tumor starts from the remaining G1 cells. A mean life span of the necrotic cells of 1.9 hr is derived from the model.