Compensatory renal growth (CRG) consists of cellular enlargement and a small but consistent increase in DNA content. It has been assumed that the increase in total renal DNA content was due to new cell formation, however, the possibility of nuclear polyploidy remained an alternative explanation. To test the hypothesis whether cellular hyperplasia is the cause of the increase in DNA content during compensatory growth after renal deprivation, we performed cell cycle analysis using flow cytometry. Following unilateral nephrectomy, the amount of cortical cells in the S phase increased by 12% at 10 h while the number of cells in the G2M phase increased by 7% at 120 h. Medullary cells entering the S phase increased by 26% at 24 h and those in G2M increased by 12% at 168 h. DNA synthesis and replication occurs during CRG following unilateral nephrectomy as evidenced by an increase in cells entering both the S and G2M phases of the cell cycle. The increase in DNA content during CRG is a result of cellular proliferation and not polyploidy.