Separation methods for glycoprotein analysis and preparation.

Abstract

Several chromatographic methods have been developed for the isolation and characterization of glycoproteins. In these methods, affinity chromatography, a single-step method, or combined use with general chromatographic methods have now become essential for the purification of many biologically important glycoproteins, including alpha1-acid glycoprotein, immunoglobulins, ceruloplasmin and erythropoietin. On the other hand, almost all glycoproteins exhibit polymorphism associated with their glycan moieties. This feature is wide-spread and has been observed in natural as well as in recombinant DNA glycoproteins. Recently, several sophisticated techniques--such as electromigration method (high-performance capillary electrophoresis) and chromatographic methods (two-dimensional polyacrylamide gel electrophoresis, high-pH anion-exchange chromatography with pulsed-amperometric detection)--have been introduced for qualitative or quantitative estimation of the microheterogeneity of glycoproteins. For gaining further insight into the structure-function relations for microheterogeneity, preparative chromatographic techniques that can yield sufficient quantities of glycoprotein variants must be developed.