Localization of isoketal adducts in vivo using a single-chain antibody.

Davies SS, Talati M, Wang X, Mernaugh RL, Amarnath V, Fessel J, Meyrick BO, Sheller J, Roberts LJ
Free Radic Biol Med. 2004 36 (9): 1163-74

PMID: 15082070 · DOI:10.1016/j.freeradbiomed.2004.02.014

Isoketals are highly reactive gamma-ketoaldehydes formed by the oxidation of arachidonic acid that rapidly adduct to proteins. To investigate the formation of isoketal adducts in vivo, we isolated and characterized a single-chain antibody from a phage displayed recombinant ScFv library that bound a model peptide adducted with synthetic 15-E2-isoketal. Recognition of isoketal adduct by this anti-isoketal adduct single-chain antibody was essentially independent of the amino acid sequence of adducted peptides or proteins. The antibody did not cross-react with 4-hydroxynonenal or 4-oxononanal adducts or with 15-F2t-isoprostane (8-iso-prostaglandin F2alpha). We investigated the formation of isoketal adducts in a well-established model of oxidative injury, hyperoxia. Exposure to >98% oxygen for 7 h dramatically increased both the number of immunoreactive airway epithelial cells and the intensity of immunoreactivity compared with animals exposed to normal room air (21% oxygen). We conclude that isoketal adducts form in epithelial cells as a result of high oxygen exposure and that this single-chain antibody provides a valuable tool to localize the formation of isoketal adducts in tissues in vivo.

MeSH Terms (20)

Aldehydes Animals Antibody Specificity Epithelial Cells Epitope Mapping Female Hyperoxia Immunochemistry Immunoglobulin Variable Region Lipid Peroxidation Lung Mice Mice, Inbred C57BL Molecular Structure Peptide Library Peptides Prostaglandins Prostaglandins E Proteins Recombinant Proteins

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