Electrotransfer of [32P]NAD allows labeling of ADP-ribosylated proteins in intact Chinese hamster ovary cells.

CHO-K1D cells electroporated in buffers containing [32P]NAD incorporated the label in a voltage-dependent manner. Electroporation with 650 V/cm at 1460 microF in Ham's F12 medium supplemented with 10 mM Hepes, pH 7.1, resulted in a greater than 20-fold increase in [32P]NAD uptake, while decreasing relative cellular survival by only 6%. Exposure of cells to gamma irradiation (20 Gy) prior to electroporation increased the steady-state level of poly(ADP-ribosylated) nuclear proteins two- to four-fold over that of unirradiated control cells. These data indicate that electrotransfer of [32P]NAD is a simple and rapid means of labeling the cellular NAD pool and should prove useful in the analysis of the relationship between poly(ADP-ribosylation) of nuclear proteins and DNA repair.