A filter immunobinding technique for the rapid detection and simultaneous identification of avian and bovine mycoplasmas

A filter immunobinding (FIB) method was developed for the detection and identification of mycoplasmas. Type strains of a total of 18 avian and bovine mycoplasma species propagated in broth media were diluted and immobilized on a nitrocellulose membrane as antigens for investigating the specificity with rabbit hyperimmune sera. Non-specific FIB reactions were easily eliminated by the procedure of absorbing rabbit hyperimmune sera in the broth. Absorbed rabbit hyperimmune sera exhibited clear species-specificity with mycoplasma antigens by the FIB. These specific reactions always agreed with the results of identification by tests of biochemical properties and growth inhibition for the isolates of M. bovirhinis, M. bovis, M. columbinum, M. columborale, M. gallisepticum and M. synoviae. Some bovine mycoplasma species, which were impossible to identify by growth inhibition test, because of their strong production of film and spots on the agar, specifically reacted with absorbed rabbit hyperimmune sera against M. bovis in the FIB. The detection limit of mycoplasmas by this method was about 104-105 colony-forming units/ml, which is lower than that of colony determination on agar. The FIB seems to be a useful technique for rapid detection and simultaneous identification of mycoplasmas.