Immunochemical analysis was employed to investigate the cell cycle-dependent protein-DNA crosslinking by cis-diamminedichloroplatinum II (cis-DDP), in HeLa-S3 cells. Cells synchronized by double thymidine block or hydroxyurea were released into S phase and incubated at 2-h intervals with cis-DDP as they progressed through S1, G2, M, and then into G1 and S phases of the subsequent cycle. Immunoblots of the DNA-crosslinked antigens reacted with antisera to 0.35 M NaCl extract or residue of HeLa S-phase nuclei revealed that several antigens changed their DNA-crosslinking pattern during the progression of HeLa cells through their reproductive cycle.