A fluorescence plate reader assay for monitoring the susceptibility of biological samples to lipid peroxidation

The susceptibility of biological samples to lipid peroxidation can be deter mined by exposing samples to a lipid peroxidation initiator and measuring t he length of time prior to the onset of lipid peroxidation. Previous studie s have shown that aldehydes generated by lipid peroxidation can react with amines to produce fluorescent products. We have utilized this principle to develop a fluorescence plate reader assay for measuring susceptibility to l ipid peroxidation. In this assay, samples are placed in glycine/phosphate b uffer and loaded into a 96-well plate. Lipid peroxidation initiators are ad ded, and fluorescence is monitored over time. Samples were assayed for susc eptibility to lipid peroxidation by both the thiobarbituric acid reactive s ubstances assay and the fluorescence plate reader assay. We found good agre ement between these two methods in assessing relative susceptibility to lip id peroxidation in liver microsomes and mitochondria. The fluorescence assa y was also used to monitor lipid peroxidation in liposomes and rat liver ho mogenates. Fluorescence was stable over an extended time period and could b e induced by a variety of lipid peroxidation initiators. The fluorescence p late reader assay offers a rapid method for monitoring lipid peroxidation i n a large number of samples. (C) 1998 Academic Press.