4-Hydroxy-2-nonenal-Trapping ELISA: Direct Evidence for the Release of a Cytotoxic Aldehyde from Oxidized Low Density Lipoproteins

Abstract

Among the aldehydes that originate from the peroxidation of cellular membrane lipids, 4-hydroxy-2-nonenal (HNE) is thought to be largely responsible for cytopathological effects observed during oxidative stress. Taking advantage of the fact that HNE is very reactive with proteins and forms stable Michael addition-type adducts, a novel immunochemical procedure for quantifying "free" HNE has been developed. The method designated as "HNE-trapping ELISA" is based on the detection of HNE trapped by a protein that has been coated in the immunoplate. The HNE-derived epitopes generated in the coating protein are then detected by the ELISA using a monoclonal antibody (mAbHNEJ-2) specific to the haptenic groups of the HNE-protein conjugates. Using this method, we determined that a considerable amount of HNE was released from human plasma low density lipoproteins (LDL) treated with copper ions or endothelial cells.