Enzyme-linked immunosorbent assay of antibody to group A Streptococcus-specific C carbohydrate with trypsin-pronase-treated whole cells as antigen

Abstract

We describe the measurement by enzyme-linked immunosorbent assay of antibody to group A Streptococcus C carbohydrate in immunized rabbits and human sera, with trypsin-pronase-treated group A streptococcal whole cells used as the antigen. The optimal concentration of the enzyme-treated whole cells used to coat the wells was 2 x 10(7) cells per well. Rabbit antiserum diluted to 1:12,800 and human serum diluted to 1:1,000 were found to be the optimal concentrations for antibody measurement. Antibody that reacted with enzyme-treated whole cells in rabbit antiserum was absorbed with group A streptococcal whole cells, purified C carbohydrate, and N-acetylglucosamine only. Enzyme-treated whole cells did not react with anti-lipoteichoic acid antibody, and rabbit antiserum did not react with lipoteichoic acid. There was a highly significant correlation between the anti-C carbohydrate antibody titrated with enzyme-treated whole cells and that with purified C carbohydrate as antigen. The correlation coefficient for the immunoglobulin M (IgM) antibodies was r = 0.75, and for the IgG antibodies it was r = 0.77. When the IgG antibody titers to the enzyme-treated whole cells of the sera of patients with acute poststreptococcal glomerulonephritis and rheumatic fever were compared with those of sera of healthy individuals, the sera of patients with poststreptococcal sequelae had significantly higher titers than did healthy individuals. Although anti-C carbohydrate antibody in human sera mostly belonged to the IgG2 subclass, there was anti-C carbohydrate antibody that belonged to the IgG3 subclass in a certain percentage of patients with rheumatic fever and acute poststreptococcal glomerulonephritis.