Immunochemistry of Salmonella O-Antigens: Specificity of Rabbit Antibodies against the O-Antigen 4 Determinant Elicited by Whole Bacteria and O-Antigen 4 Specific Saccharide-Protein Conjugates

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Abstract
In two precipitating systems using the O-antigenic polysaccharide (PS) from Salmonella typhimurium as antigen and BO antiserum (anti-O4 and -O12 specificities) or O4 antiserum, the octasaccharide was the best inhibitor. In order of decreasing effectiveness followed: . On a molar basis the relative inhibiting activities were 100:45:12.5:0.3:0.1 (BO antiserum) and 100:40:8:0.3:0.1 (O4 antiserum). These data established that the combining sites of antibodies raised by the native bacterial O4 antigen recognize a structure larger than the tetrasacharide but equal to or smaller than the octasaccharide. When the S. typhimurium PS was precipitated with antiserum elicited by immunization with the octasaccharide (see above) conjugated to bovine serum albumin (Os-BSA) the relative inhibitory activities of four of the saccharides (not including methyl-β-abequoside) were 100:30:1.6:0.1. Thus, the combining sites of antibodies elicited by the Os-BSA conjugate recognized structures close or equal to those of antibodies raised by whole bacteria (BO and O4 antisera). When the S. typhimurium PS was precipitated with an antiserum elicited by immunization with 3-O-α-abequopyranosyl-D-mannopyranose covalently linked to BSA (AM-BSA) the relative inhibitory activities of the five saccharides were 7.5:17:100:0.6:0.2. Hence, the AM-BSA antibodies had about fifteen times higher affinity for the methyl 3-O-α-abequopyranosyl-α-D-mannopyranoside than for the octasaccharide. The importance of using different sized haptens, e.g., di- or octasaccharides, to elicit antibodies useful for diagnostic or immunoprophylactic purposes is emphasized.