Corneal glycosaminoglycan synthesis in long-term organ culture.
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Biomedical subjects
J L Funderburgh
Publications and source records attributed to J L Funderburgh.
An agarose gel electrophoretic method for analysis of sulfated glycosaminoglycans of cultured cells.
Agarose disc gel electrophoresis has been adapted to achieve the separation of the major sulfated glycosaminoglycans produced by cells in culture. By use of buffers containing barium ion, mixtures of chondroitin sulfate, dermatan sulfate, and heparan sulfate are well resolved into discrete bands. The technique can be used preparatively as well as analytically to separate quantities of glycosaminoglycans up to a milligram in a 6-mm diameter gel.
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Monoclonal antibodies to rabbit corneal keratan sulfate proteoglycan.
Six mouse hybrid cell lines were isolated which secrete antibodies to rabbit corneal proteoglycan. All six antibodies interacted with the same fraction of the proteoglycan, precipitating approximately 50% of proteoglycan labelled in the protein moiety. A radioimmunoassay using these antibodies measured concentrations as low as 1 g/ml unlabelled rabbit corneal proteoglycan. Human corneal proteoglycan, corneal keratan sulfate, and an oligosaccharide fraction from corneal digests all interacted with the antibodies at concentrations similar to whole rabbit proteoglycan. Proteoglycans from cultured rabbit stromal fibroblasts and from sclera were 20 to 50-fold less effective in competition for antibody. Endo-beta-galactosidase treatment of proteoglycan reduced antibody binding, but protease or chondroitinase treatments did not. Labelled proteoglycan separated by antibody affinity chromatography contained only keratan sulfate, whereas proteoglycan not bound to affinity columns contained only chondroitin sulfate. The antibodies appear to recognise a carbohydrate structure found only on corneal keratan sulfate proteoglycan. This structure can serve as a basis for separation. This structure can serve as a basis for separation of corneal proteoglycan types using antibody affinity chromatography.
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