Structural characterization of glycoprotein carbohydrate chains by using diagoxigenin-labeled lectins on blots

Abstract

The carbohydrate structures of blotted glycoproteins can be analyzed by probing them with lectins. Here we describe a method where lectins conjugated with digoxigenin are used in combination with an anti-digoxigenin antibody AP conjugate as a very sensitive detection system for this type of analysis. The specificity of the lectins used, and the sensitivity of the detection system, provide valuable conclusions on the glycan structures. Only small amounts of glycoproteins are required for the analysis. The binding specificity of a set of lectins is demonstrated with various glycoproteins of defined carbohydrate structure. The application of these labeled lectins in combination with specific glycosidases for the characterization of the carbohydrate chains of recombinant tissue plasminogen activator and erythropoietin is presented.