cPLA2 Assay Kit Usage And Synthesis

Description

Phospholipase A2 catalyzes the hydrolysis of fatty acids at the sn-2 position of glycerophospholipids, yielding a free fatty acid and a lysophospholipid as products.1 The release of arachidonic acid from membrane phospholipids by these enzymes is believed to be the key step in the biosynthesis of eicosanoids.2 There are primarily three different kinds of phospholipase A2. They are secretory (sPLA2), calcium-dependent cytosolic (cPLA2), and calcium-independent cytosolic (iPLA2) phospholipase A2. Of these three different types of enzymes, only the cPLA2 exhibits specificity towards arachidonic acid whereas all others can hydrolyze any fatty acid at the sn-2 position. Arachidonoyl Thio-PC is a substrate for cPLA2 by virtue of the presence of arachidonic acid at the sn-2 position of the glycerophospholipid.3 Hydrolysis of the arachidonoyl thioester bond at the sn-2 position by PLA2 releases a free thiol which can be detected by DTNB (5,5'-dithio-bis-(2-nitrobenzoic acid)). This assay can be used to determine the activity of cPLA2 in purified preparations, cell cultures, or tissue homogenates that are known to contain only cPLA2. Use of this assay with preparations containing more than one type of PLA2 will result in the measurement of total PLA2 activity rather than cPLA2 alone. Isozyme-specific cPLA2 activity can be measured by excluding sPLA2 or inhibiting iPLA2 activities in the assay. Each kit contains cPLA2 assay buffer, DTNB/EGTA, Arachidonoyl Thio-PC (substrate), bee venom PLA2 (control), bromoenol lactone solution, a 96 well plate, and complete instructions.Needed but not supplied: Please download the kit booklet to verify if UltraPure Water (Milli-Q or equivalent) or any other components are needed for this assay.WARNING This product is not for human or veterinary use.