Mice immunised with partially purified surface molecules of photoreceptor outer segments or with a synthetic peptide corresponding to the N-terminus of a cone-specific rhodopsin-like protein, gave rise to the production of monoclonal antibodies with only general immunoreactivity against the retina. Immunisation with interphotoreceptor matrix or crude photoreceptor outer segment preparations resulted in the production of hybridomas which secreted monoclonal antibodies 1001.A1 and 1001.A3. 1001.A1 binds to the interstitial retinol-binding protein (IRBP), associated with the rod photoreceptor cells and the IPM, as demonstrated by immunohisto chemistry and by Western blotting and dot blotting of IPM and purified IRBP. 1001.A3 binds a soluble high molecular weight chondroitin sulphase proteoglycan. Immunohistochemistry indicated the antigen to be present in the form of distinct sheath-like structures surrounding the photoreceptor cells. Gel filtration chromatography both in native conditions and in the presence of guanidinium chloride showed the antigen to have an apparent molecular mass of greater than 2000kDaltons and indicated the antigen was not a loosely associated aggregate of smaller components. Binding of 1001.A3 to fixed tissue sections of the bovine retina was completely abolished by their prior treatment with either chondroitinase ABC, chondroitinase AC, hyaluronidase (testicular) or trypsin. Prior treatment of tissue sections with either heparinase or neuraminidase had no effect on binding. Treatment of tissue sections with hyaluronidase (<i>Streptomyces</i>) had no effect on the ability of 1001.A3 to bind, but the structure of the antigen was altered. The sheath-like structure surrounding the photoreceptors was broken down and immunoreactivity was seen in the same area of the IPM, adjacent to the photoreceptors but with no defined structure. In conclusion, the antigen is a chondroitin sulphate molecule which is associated with hyaluronic acid molecules and which together form a defined sheath-like structure surrounding the photoreceptor cells.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:661009 |
| Date | January 1989 |
| Creators | Reid, K. |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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