Using multiparameter flow cytometric analysis, we find that senescent cells accumulate in a unique cell-cycle compartment characterized in cell-cycle arrest in G1 and a significantly reduced nucleocytoplasmic ratio (genome size/cell mass) relative to cycling cells. With respect to gross cellular phenotype, the quiescent state of senescent cells differs from quiescence induced by density inhibition; the former is associated with a reduction in the nucleocytoplasmic ratio, while the latter is associated with an increase in the nucleocytoplasmic ratio. Senescent cells were present at all passages examined. The frequency of senescent cells was low in early-passage cultures and increased with passage number. Senescence of populations of IMR-90 cells reflects change in the relative frequency of these cells. The frequency of cells with karyotypic changes increased with the progressive accumulation of out-of-cycle cells.