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Show posters on the hill Novel Assay of Phospholipase A2 Enzymatic Activity using Electrospray-Mass Spectrometry Grant Myers*, Joel Harris** Departments of Bioengineering* and Chemistry** I.'.'"., I ,..,.,,'. lhcchmdemlit: a:\titm-: I'im :¦;.>:;¦.:.. \ .• ¦ Pt A2 has bci.il linked to tissue damage ifi rheumatoid arthritis, mvoeardi and acute pancreatitis. Researcher* have proposed the Chloroform Conhol Bath w Measuring the Reaction Rate i novel procedua- was adapted u> ;mal> /* Hk rale t>f the en/> me. Pi .A2 U sddcO to les and limed sample* are pcrioditalh rctiHivctt. f'hi>sp!K)lipiits arc recovered bj i.r»i di^Kcd in d)^.M»l^.n-mcthan..i i\i I-.SI-MS The Mi.innu,. tjiiattro II Instrumentation In Uectrosprav loni/.nion M.iss S|vcttontetr> <! SI-MM. evaponttion of solvent and theeleelroslatie repulsion ulttiur^ed ilropleis resnl!, in flit iiuii/mioti of sample molecules to allow mass spccliomi-m iin.iKsis \\liitli generates a spectrum of ionk in the Novel Assay of Phospholipase A2 Enzymatic Activity Using Electrospray-Mass Spectrometry Grant Myers, Joel Harris Departments of Bioengineering and Chemistry X hospholipids comprise the primary molecular structure in cell membranes, and their chemical stability assures the integrity of the membrane and the viability of the cell. Phospholipase A2 (PLA2) is a relatively common enzyme that catalyzes the hydrolysis of phospho-lipids leading to destruction of cell membranes. The products released by this reaction are key inflamma- complexity to the overall action and kinetics of the enzyme. Traditional assay methods for this enzyme such as pH-stat, fluorescence, and radiolabel-ing are time consuming, use expensive reagents, and require high lipid concentrations. Electrospray- Ionization Mass Spectrometry (ESIMS) has excellent sensitivity, utilizes common laboratory chemicals and requires only small volumes and concentrations of lipids. ESI-MS uses 'soft' ionization to separate sample components by mass and charge, leaving ionized molecules intact which simplifies quantitative interpretation of the results. A protocol developed for ESI-MS analysis of lipids in this research utilizes extraction techniques developed in 1959 for the gravimetric analysis of high concentrations of lipids in frozen fish. This protocol was adapted to concentrations of lipids in small volume samples and allows the chemically selective investigation ofPLA2 reaction kinetics using ESI-MS. tory signals used by cells of the immune system. Uncontrolled PLA2 has been linked to tissue damage in inflammatory syndromes such as Rheumatoid Arthritis, Acute Pancreatits, Myocardial Infarction, Asthma, Septic Shock and allergic reactions. In contrast to many enzymes, PLA2 preferentially acts on lipids that are organized into membranes and not isolated lipid molecules. This fact increases the |