Effect of duration of incubation ofMycoplasma gallisepticumcultures on the sensitivity and specificity of antigens for ELISA and microimmunofluorescence tests

Abstract

In the production of Mycoplasma gallisepticum (Mg) antigens for ELISA and microimmunofluorescence (MIF) tests, two strains of Mg, S6 and PG31, were grown in broth culture and harvested at intervals from 18 to 138 h. The effect of duration of incubation of culture on antigen sensitivity and specificity was assessed using homologous sera (against Mg S6), and sera prepared against Mycoplasma synoviae (Ms), and against mycoplasma media. It was found, in both Mg S6 and PG31 ELISAs, that in the period 18 to 70 h of incubation, sensitivity declined with homologous sera, but little thereafter. A more rapid and greater decline occurred with PG31 antigen than S6. The antigens showed a lack of specificity since antisera to Ms gave positive reactions. These were not influenced by duration of incubation of Mg culture. In the Mg S6 ELISA lack of specificity was also shown by positive reactions with sera against media components. These increased in the first 42 h of cultivation. In the MIF test there was no loss of sensitivity with Mg S6 antigen with increasing duration of incubation of culture but it occurred with PG31 antigen. The non-specific reactions with antisera to Ms increased with increased duration of incubation of both strains of Mg. This work demonstrates that the most sensitive antigens are produced by harvesting organisms early in culture, although this does not eliminate the Ms cross reactions.