Enzyme-linked immunosorbent assay for human IgG, IgA, and IgM antibodies to antigens from anaerobic cultures of seven oral bacteria.

Monocultures of 7 oral bacteria were grown anaerobically, and antigens were partially purified from the supernatant fluids by gel filtration. Isoelectric focussing showed that the antigen fractions contained PAS-positive material and proteins focussing between pH 3.5 and pH 5.5. Specific activity against the 7 antigen fractions was observed for IgG, IgA, and IgM in serum from patients with periodontal disease by means of enzyme-linked immunosorbent assay (ELISA). Conditions for antigen coating in ELISA were studied, and large variations with regard to optimal concentration were found among the 7 fractions. The ELISA readings were virtually unaffected by increasing the pH of the coating buffer from 5 to 9. Addition of sheep serum to the incubation buffer was disadvantageous as the serum showed moderate antibody activity to the 7 antigen fractions, and human serum apparently contained IgM antibodies to sheep IgG. Addition of 0.5% BSA improved the reproducibility of ELISA considerably, although the sensitivity was somewhat decreased.