Isolation and partial characterization of the heterophile antigen of infectious mononucleosis from bovine erythrocytes

Abstract

The heterophile antigen (Paul‐Bunnell antigen, PBA) of infectious mononucleosis was isolated by extraction of an aqueous suspension of bovine erythrocyte stromata with chloroform‐methanol (2:1). The upper aqueous layer contained gangliosides, PBA, and a high‐molecular‐weight glycoprotein. PBA and gangliosides were separated from the high‐molecular‐weight glycoprotein by extraction of lyophilized upper layer with chloroform‐methanol solvents. Separation of PBA from gangliosides was carried out by chromatography on DEAE‐cellulose with chloroform‐methanol solvents. PBA appeared to be a minor glycoprotein component of the erythrocyte membrane and had both hydrophobic and hydrophilic properties. It was soluble in either organic or aqueous solvents. On SDS‐polyacrylamide gel electrophoresis, it migrated as a single component that stained for protein with Coomassie blue, for carbohydrate with periodic acid‐Schiff reagent, and for lipid with oil red 0; it had an apparent molecular weight of 26,000. It was composed of 62% protein with major amino acids: glutamic acid, proline, glycine, isoleucine, leucine, and threonine (158, 116, 98, 90, 85, and 82 residues per 1,000 residues, respectively). Carbohydrate content was 9.2% with major sugar constituents: sialic acid, galactosamine, and galactose. Serologic activity of PBA was destroyed by pronase but not by trypsin.