Phospholipase A2 and Phosphatidylinositol-Specific Phospholipase C Assays by HPLC and TLC with Fluorescent Substrate

Abstract

Lipolytic enzymes have traditionally been assayed by radiometric and titrimetric methods (1). Radiometric methods are quite sensitive but require expensive radiolabeled substrates and tedious separation of labeled substrate and products. In addition, the safe use of radioactive materials is of increasing concern. Titrimetric assays are continuous and quite straightforward and use natural substrates but suffer from low sensitivity and are subject to conditions that may alter the amount of free hydrogen ions released. Fluorescence-based assays have sensitivities that approach those of radiometric methods; although they require synthetic fluorescent-labeled substrates, they are often more convenient and rapid. For a recent review, see Hendrickson (2).