Production and Specificity of Antibodies Against the Central Region of Type Ii Collagen

Abstract

Collagen type-specific antibodies as well as antibodies to particular portions of the molecule are extremely useful tools especially for the quantification of collagens and for immunohistochemical examinations in developing embryos. Quantification of collagens in CNBr-solubilized tissue samples presupposes the production of antibodies against CNBr-derived collagen fragments. For the first time, as antigens for the immunization of rabbits, cyanogen-bromide derived fragments of collagen type II were used, obtained by direct digestion of tissue (Swarm chondrosarcoma from rat) and separation by gel filtration chromatography. Antisera were applied to affinity chromatography and the eluted antibodies were characterized by ELISA, immunoblotting, inhibition studies and immunohistochemistry. The antibodies from five different rabbits show high specificity for type II collagen and are directed against sequential determinants in the central portion of the type II collagen molecule. The easy way of obtaining the fragments directly from tissue, combined with their immune response in rabbits, gives the possibility of producing type II collagen-specific, fragment-directed antibodies in a convenient and rapid way.