ISOLATION OF OLIGOSACCHARIDES--SUGAR-BINDING PROTEINS

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Carbohydrates have been implicated in many physiologically important cell surface functions including cell-cell recognition, growth control and receptors for toxins and glycoprotein hormones. The difficulties in obtaining sufficient amounts and determining the structure of the limited quantities of physiologically important cell surface oligosaccharides are major reasons for the lack of knowledge about what structures are involved in these processes. Since free oligosaccharides in human milk contain structures also found in cell surface glycolipids, antibodies against these oligosaccharides can be used in sensitive radioimmunoassays to detect specific carbohydrates in complex biological samples. Since chemical methods for structural analyses of abnormal oligosaccharides, diagnostic for certain diseases and genetic disorders of their metabolism, require laborious purification and expensive instrumentation, they are generally not applicable to screening, diagnosis or classification of diseases. Radioimmunoassays for diagnostically important oligosaccharides may be a valuable alternative. As a basis for this application, specific antisera against human oligosaccharides will be applied to the ganglioside analysis of human meconium, a potential diagnostic specimen and a rich source of fetal glycolipids. Cell surface glycolipids that bind 125I-labeled carbohydrate-specific antibodies, cholera toxin, lectins and other physiologically important carbohydrate-binding proteins can be detected on thin layer chromatograms by sensitive autoradiographic techniques. Oligosaccharides released from functional glycolipids can be labeled by reduction with NaB[3H]4 and isolated from complex mixtures by radioimmune-binding methods. Structural analyses using specific anti-oligosaccharide antibodies and specific exoglycosidase digestion will be fundamental to identifying minor, functional cell surface carbohydrate structures. These methods will also be applied to the characterization of blood group specific lectins and antibodies and their corresponding glycolipid isoantigens.