Abstract
Hepatitis B surface antigen (HB sAg) was isolated from human serum by two steps of affinity chromatography on antibody-coated gels. HB sAg-positive serum was passed through a column packed with guinea pig anti-HB sAg antibodies covalently bound to CNBr-activated beaded agarose gel. The majority of non-specifically bound protein was removed by washing the gel with increased concentrations (0.5 M) of NaCl in Tris buffer. Elution of the specifically bound HB sAg was carried out with 3 M NaSCN. Residual normal human serum proteins present in the eluate were removed by passing the partially purified HB sAg 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 haemagglutination in the isolated HB sAg. Cross-reactions between HB sAg 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.
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