A highly specific two-site ELISA for pneumococcal C-polysaccharide using monoclonal and affinity-purified polyclonal antibodies.
A two-site ELISA for the detection of pneumococcal C-polysaccharide (PnC) has been developed. A monoclonal antibody directed against the phosphorylcholine residue of the PnC was used as catcher and an affinity-purified polyclonal anti-PnC rabbit antiserum for detection. Polyclonal antibodies against the PnC as well as capsular antigens were obtained by immunizing rabbits with type 1 pneumococci. Antibodies against the phosphorylcholine determinant of PnC could be removed by affinity purification. Remaining antibodies reacted in an ELISA with type 1 capsular polysaccharide as well as with PnC. Only in the fraction with the highest antibody activity against PnC, phosphorylcholine exhibited a slight inhibitory action. It is concluded that the purified antibody preparation reacted with an antigenic determinant shared by the two polysaccharides, in all probability a determinant associated with 2-acetamido-4-amino-2,4,6-trideoxygalactose which is the only monosaccharide component in common between PnC and the type 1 capsular polysaccharide. By the use of this affinity-purified antibody preparation, reactions with alpha-streptococci, occurring with non-purified serum, were abolished. The sensitivity and specificity of the test was determined using capsulated and non-capsulated pneumococci and alpha-streptococci known to cross-react with unpurified serum against the pneumococcal C-polysaccharide.
D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology* / Q000302:isolation & purification; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000942:Antigens, Bacterial / Q000032:analysis*; D002846:Chromatography, Affinity; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D011135:Polysaccharides, Bacterial / Q000032:analysis*; D011817:Rabbits
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D011135:Polysaccharides, Bacterial; C027158:polysaccharide C-substance (Streptococcus)