Antigens from trichinella spiralis that induce a protective response in the mouse

Abstract

A series of monoclonal antibodies was generated for the purpose of studying antigens of the infective L1 larva of Trichinella spiralis. Primary immunization of donor BALB/c mice was by oral infection with L1 larvae. Secondary immunization and screening procedures for hybridoma clones employed a pool of affinity-purified antigens recognized by pooled serum from infected rabbits. Three monoclonal antibodies, chosen for their high reactivity in screening assays and their unique specificities, were raised in quantity, purified, and attached to Sepharose 4B for affinity chromatography. The antigens selected by each affinity column were of different m.w. (37K, 48K, and 50/55K) and possessed different abilities to induce protection. The 48K antigen induced a high level of protection at biologically relevant doses (1.0 and 0.1 micrograms protein/mouse). The 50/55K antigen induced a lower level of protection than the 48K antigen, but was still effective at similar doses. The 37K antigen elicited protection only at a dose of 50 micrograms protein/mouse, and was not as effective as the unpurified antigen at any dose. Earlier studies indicated that many antigens involved in the immune response are secreted. Therefore, worm secretions and two subcellular fractions of homogenized worms (S2 = cytosol, S3 = soluble portion of the large particle fraction) were analyzed for content of the three purified antigens by a double antibody sandwich enzyme-linked assay. The 48K and 50/55K antigens represent significant components of the secretions (12% and 5%, respectively) but relatively minor components of the S2 and S3 fractions. The 37K antigen represents a major component of the S2 fraction, but is essentially absent from S3 and secretions. Immunocytolocalization studies employing immunoperoxidase methods on infected muscle tissue show that the 48K antigen is located in the beta-stichocytes, the lining of the gut, and the surface of the cuticle. The 50/55K antigen was detected in the alpha-stichocytes and was occasionally observed in the gut or on the cuticle. The 37K antigen was detected only in the pseudocoelom. The results from the quantitative ELISA and immunocytolocalization show that the two antigens that induce a protective response are secreted proteins. These data constitute the first account of antigens purified from a nematode that induce a strong protective response. One of these (48K) confers a level of protection comparable to that elicited by exposure of an entire infection.