An immunohistochemical method of detecting Mycoplasma species antigens by use of monoclonal antibodies on paraffin sections of pneumonic bovine and caprine lungs

Abstract

Lung samples from pneumonic lesions in cattle and goats, naturally or experimentally infected with strains of the Mycoplasma mycoides cluster, were fixed in formalin and embedded in paraffin. An immunohistochemical technique using monoclonal or polyclonal antibodies was performed on tissue sections in order to detect Mycoplasma antigens. Four monoclonal antibodies (MAbs), one (2A3) raised against M. mycoides ssp. mycoides small colony (SC) and large colony (LC), two (1D3 and 5E5) against M. mycoides ssp. capri, and one (5A10) against M. bovis, were used. A range of polyclonal antibodies, raised to the individual subspecies of the M. mycoides cluster, and one to Pasteurella haemolytica, was also used. The MAb 2A3 showed positive immunostaining in lung sections from cattle and goats naturally and experimentally infected with M. mycoides ssp. mycoides SC and LC, but not with pneumonic lesions of cattle and goats due to other members of the M. mycoides cluster, M. bovis or Pasteurella spp. The MAb 1D3 showed immunostaining in lung sections from goats naturally and experimentally infected with M. mycoides ssp. capri, but again not with pneumonic lesions caused by other members of the M. mycoides cluster, M. bovis or Pasteurella spp. The MAb 5E5 immunoreacted in sections from pneumonic lesions from all animals infected with one of the three M. mycoides cluster subspecies used in the study, but not with M. bovis or Pasteurella infected tissue. Immunoreaction was mainly found in the cell debris around necrotic areas, as well as in macrophages, neutrophils and epithelial cells. The localization of antigens of the M. mycoides cluster using polyclonal antisera followed basically the same pattern as that obtained with the monoclonals. However, a wide cross reactivity was found between different antisera and relatively high background immunostaining was also seen, especially in necrotic areas. The results suggest that immunohistochemical methods using monoclonal antibodies are useful tools for the diagnosis and study of the pathogenesis of pneumonia caused by the Mycoplasmas of the M. mycoides cluster.