title: A monoclonal antibody recognizing the chlorohydrin derivatives of oleic acid for probing hypochlorous acid involvement in tissue injury.
abstract: A monoclonal antibody against hypochlorous acid-modified oleic acid has been raised to investigate involvement of HOCI in tissue injury. Mice were immunized with an isomeric mixture of chlorohydrin derivatives of oleic acid (18:0-chlorohydrin) conjugated to keyhole limpet haemocyanin (CH-KLH). The chlorohydrin was formed by the treatment of oleic acid with hypochlorous acid. Monoclonal antibodies were raised and the fusion was screened with 18:0-chlorohydrin-bovine serum albumin (CH-BSA) conjugate. A number of antibody-secreting clones were identified and the supernatants were characterized by binding studies and dose-response curves. In ELISA, mAb CH-1 had an equivalent titre when either the chlorohydrin or bromohydrin derivative of oleic acid, complexed to bovine serum albumin, was used as screening antigen. The mAb CH-1 recognition of CH-BSA was competed with chlorohydrin and bromohydrin conjugates of BSA and KLH. Similarly, free 18:0-chlorohydrin and the 18:0-chlorohydrin-phosphatidyl choline treated with hypochlorous acid competed with mAb CH-1 binding. The mAb CH-1 also recognised the chlorohydrin derivative of linoleic acid and chlorohydrin formed from palmitoyl, oleyl phosphatidyl choline but with a decreased avidity. Weak cross-reactivity was observed with hydroxy-linoleic acid and linoleic acid hdroperoxide, either as free fatty acid or in phosphatidyl choline. There was minimal competitive binding of mAb CH-1 to free oleic acid, 16:0/18:1 phosphatidylcholine, cholesterol, or cholesterol chlorohydrin. The mAb CH-1 described here may be a useful probe for assessing the involvement of hypochlorous acid in tissue injury.
authors.affiliation: a Department of Pathology , Christchurch School of Medicine , P.O. Box 4345, Christchurch , New Zealand.
authors.forename: N M
authors.initials: NM
authors.lastname: Domigan
authors.affiliation: a Department of Pathology , Christchurch School of Medicine , P.O. Box 4345, Christchurch , New Zealand.
authors.forename: A C
authors.initials: AC
authors.lastname: Carr
authors.affiliation: b Steroid & Immunobiochemistry Laboratory, Clinical Biochemistry Unit, Canterbury Health Laboratories , Christchurch , New Zealand.
authors.forename: J G
authors.initials: JG
authors.lastname: Lewis
authors.affiliation: b Steroid & Immunobiochemistry Laboratory, Clinical Biochemistry Unit, Canterbury Health Laboratories , Christchurch , New Zealand.
authors.forename: P A
authors.initials: PA
authors.lastname: Elder
authors.affiliation: a Department of Pathology , Christchurch School of Medicine , P.O. Box 4345, Christchurch , New Zealand.
authors.forename: C C
authors.initials: CC
authors.lastname: Winterbourn