Antigenic and Polypeptide Structure of Turkey Enteric Coronaviruses as Defined by Monoclonal Antibodies

Abstract

Twenty-nine hybridoma cells lines, producing monoclonal antibodies (MAbs) to the Minnesota strain of turkey enteric coronavirus (TCV), have been established by fusion of Sp2/0 myeloma cells with spleen cells from BALB/c mice immunized with purified preparations of the egg-adapted or tissue culture-adapted virus. The hybridomas produced mainly IgG2a or IgG1 antibodies. Western immunoblotting experiments with purified virus, and immunoprecipitation tests with [35S]methionine-labelled infected cell extracts, allowed assessment of the polypeptide specificity of the MAbs. Sixteen hybridomas secreted antibodies directed to the peplomeric protein (E2, gp200/gp100) and putative intracellular precursors of apparent Mr 170K to 180K and 90K. Four hybridomas produced antibodies that selectively reacted with a glycoprotein with an Mr of 140K (E3). This polypeptide species corresponded to the major structural component of small granular projections, located near the base of the larger bulbous peplomers, and was found to be responsible for haemagglutination. The major neutralization-mediating determinants were found to be carried by both E2 and E3 glycoproteins. Eight hybridomas produced MAbs directed to the major nucleocapsid protein (N, 52K), and only one MAb reacted with a low Mr structural glycoprotein (24K), corresponding to the matrix (E1) protein. By indirect immunofluorescence, MAbs of different specificity also revealed distinct patterns of distribution of the viral antigens within the cells. The location on the virion of the antigenic determinants recognized by MAbs of different specificity was determined by the use of an immunogold electron microscopy technique. Comparison of nine TCV Quebec strains, using MAbs directed to peplomer and haemagglutinin proteins of the prototype Minnesota strain, confirmed their close antigenic relationship, but also revealed the occurrence of at least two distinct antigenic groups.