A SEMIAUTOMATED 96-WELL PLATE ASSAY FOR PROTEIN KINASE-C

We have devised a rapid procedure for the assay of protein kinase C. Reactions (100 μl) were set up in the wells of a 96-well assay plate and initiated one column at a time by the addition of [γ-32P]ATP with an eightchannel pipettor. After incubation for 5 min at 30°C, the reactions were terminated by the addition of 100 μl of 25% trichloroacetic acid. The reaction mixtures were then filtered using a semiautomatic cell harvester, and transferred to scintillation vials using a filter punch apparatus. Direct comparison of this method to traditional techniques revealed a three- to fivefold increase in efficiency with equal sensitivity. This method is suitable for screening large numbers of inhibitors and activators of protein kinase C and appears to be applicable to other enzymes such as calmodulin kinases. © 1990.