The fluorescence scanning of microgels stained for glycoproteins with fluorescein isothiocyanate-labelled concanavalin A.
A technique is presented which allows one to label and quantitate glycoproteins. Small amounts of protein from biological samples (0.5-2.5 microgram for mixtures and less for individual proteins) are separated by sodium dodecyl sulfate gel electrophoresis on 1-30% polyacrylamide gradient microgels. The gels are stained with Co-omassie Brillant Blue R250 to evaluate relative migration or fixed in 2-propanol/acetic acid and stained with fluorescein isothiocyanate-labelled concanavalin A. The microgels are then scanned using a fluorescence microscope controlled by a computer, although simpler configurations are possible. Standards of known carbohydrate composition (e.g., glucose oxidase) are used for comparative purposes. Glycoproteins in the order of 5-30 ng protein (or 1-5 ng carbohydrate) can be detected without difficulty. This technique may prove valuable in evaluating glycoproteins when the available material is limited.
D000818:Animals; D003208:Concanavalin A; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005452:Fluoresceins; D005453:Fluorescence; D006023:Glycoproteins / Q000032:analysis*; D008970:Molecular Weight; D009186:Myelin Sheath / Q000032:analysis
D005452:Fluoresceins; D006023:Glycoproteins; D003208:Concanavalin A