A fluorogenic analog of the PLA(2) substrate PC, named Dabcyl-BODIPY-PC, or simply DBPC, was synthesized with a fluorescence quencher (Dabcyl, 4-[(4-[N,N-dimethylamino]phenyl)azo]benzoic acid) in the sn-1 acyl chain and a BODIPY fluor in the sn-2 acyl chain. DBPC was recognized by sPLA(2) from each of the four sources examined (bee venom, human synovial fluid, cobra venom, and bovine pancreas). A dramatic and quantifiable fluorescence enhancement of DBPC occurred upon phospholipase digestion both in the presence and absence of excess PC. Both real-time and endpoint assays for PLA(2) were sensitive, consistent, and rapid. Thus, DBPC can be used as a sensitive fluorogenic probe for in vitro high-throughput screening assays for PLA(2) activation and inhibition and would expedite studies of PLA(2) in cellular signaling, in vitro screening for drug discovery, and subcellular localization of enzyme activity.