Isolation of hepatitis B surface antigen (HBsAg) by affinity chromatography on antibody-coated immunoadsorbents.
Hepatitis tb surface antigen (HBsAg) was isolated from human serum by two steps of affinity chromatography on antibody-coated gels. HBsAg-positive serum was passed through a column packed with guinea pig anti-HBsAg antibodies covalently bound to CNBr-activated beaded agarose gel. The majority of non-specifically bound proteins was removed by washing the gel with increased concentrations (0.5 M) of NaCl in Tris buffer. Elution of the specifically bound HBsAg was carried out with 3 M NaSCN. Residual normal human serum proteins present in the eluate were removed by passing the partially purified HBsAg through an immunoadsorbent coated with rabbit antibodies directed against human serum proteins. After this treatment normal human serum proteins could no longer be demonstrated by passive hemagglutination in the isolated HBsAg. Cross-reactions between HBsAg and normal human serum proteins could not be demonstrated. Both antibody-coated immunoadsorbents could be used over ten times without significant loss of their binding capacity.
D000017:ABO Blood-Group System; D000327:Adsorption; D000818:Animals; D001798:Blood Proteins / Q000302:isolation & purification; D005591:Chemical Fractionation; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D003488:Cyanogen Bromide; D004912:Erythrocytes / Q000276:immunology; D006168:Guinea Pigs / Q000276:immunology; D006386:Hemagglutination Tests; D006509:Hepatitis B / Q000276:immunology*; D006510:Hepatitis B Antibodies; D006511:Hepatitis B Antigens / Q000302:isolation & purification*; D005779:Immunodiffusion; D008722:Methods; D012685:Sepharose; D013634:Tannins; D005719:gamma-Globulins
D000017:ABO Blood-Group System; D001798:Blood Proteins; D006510:Hepatitis B Antibodies; D006511:Hepatitis B Antigens; D013634:Tannins; D005719:gamma-Globulins; D012685:Sepharose; D003488:Cyanogen Bromide