Leaching of concanavalin A during affinity chromatographic isolation of cell surface glycoproteins from human fetal neurons and glial cells

Abstract Preparation of surface glycoproteins from human fetal brain cells by affinity chromatography on Con A-Sepharose 4B was a problematic endeavor due to leaching of Con A from the matrix. Dissociation of Con A from the matrix took place irrespective of the presence of lipid and/or detergent and the buffer composition during chromatography and was apparently related to the nature of the protein under study. Pretreatment of Con A-Sepharose with 6 m guanidine or 8 m urea reduced Con A leaching. The Con A eluate also contained noncovalently associated glycolipid. Elution at 25°C rendered fractions containing a higher degree of Con A and glycolipid contamination compared to the negligible contamination by these two components when elution was carried out at 4°C. This phenomenon was attributed to the formation of heterogeneous mixed micelles of glycoprotein.