To allow more sensitive, selective, and routine analyses of platinum(Pt)-GG and -AG intrastrand crosslinks we have significantly improved our quantitative 32 P-postlabeling assay (M. J. P. Welters et al. Carcinogenesis 18, 1767Carcinogenesis 18, -1774Carcinogenesis 18, , 1997)). Instead of off-line scintillation counting we introduced an on-line flow radioisotope detector into the HPLC system. Furthermore, the isolation protocol for the adducts was significantly modified and optimized to reduce interfering background peaks that prevented quantification of low levels of the cisplatin-DNA adducts in white blood cells obtained from patients. Reduction of background signals was obtained by boiling the samples, followed by phenol/chloroform/isoamylethanol extraction after the DNA digestion step. The labeling efficiency for the adducts was increased by 40% by using Na-formate instead of NH 4 -formate for elution of the adducts from the strong cation-exchange columns. Finally, a calibration curve and quality controls were implemented. The labeling efficiencies were not different between the dinucleotides. The between-and within-run precision for the Pt-GG and Pt-AG adducts measured at the lower limit of quantification of 87 and 53 amol/g DNA, respectively, was less than 20% CV. The adducts were stable in DNA stored for a 2-month time period at Ψ80Β°C. The assay is now routinely used for high-precision analyses of patient and cell line samples containing very low adduct levels.