SUMMARYWe established three hybridomas (designated 2A12, 4C4 and 6H8) secreting monoclonal antibodies which react with N and H(heated) antigenic particles of poliovirus type 1 (Mahoney strain). One of these monoclonal antibodies, 4C4, had virus-neutralizing activity, while the other two were non-neutralizing. A solid-phase radioimmunoassay and a neutralization test indicated that the specificity of the neutralizing monoclonal antibody 4C4 was the same as that of the monospecific antibody HN31, previously prepared by us by absorbing poliovirus antiserum with an equine serum inhibitor-resistant mutant (M-HN31) of poliovirus. Furthermore, by means of immune electron microscopy, it was revealed that the combining site of the 4C4 neutralizing monoclonal antibody is on or around vertices of the N and H antigenic particles, while the other two monoclonal antibodies are directed to the entire surface of the antigens.Two antigens have been found in poliovirus: the N antigen associated with infectious complete particles and the H antigen, which includes naturally occurring empty capsids and 80S particles converted from N antigen by exposure to heat, u.v. radiation, or high pH (Mayer et al., 1957;Roizman et al., 1959; Boey6 & Van Elsen, 1967, Katagiri et al., 1967, 1971Breindl, 1971). Monoclonal antibody preparations produced by hybridoma cells have proved useful for studying the antigenic structure of several antigenic components of poliovirus (Icenogle et al., 1981;Brioen et al., 1982;Emini et al., 1982;Thorpe et al., 1982;Blondel et al., 1983). Emini et al. Icenogle et al. (1981) established hybridomas producing neutralizing monoclonal antibodies that react with N antigen (145S) and, to a lesser extent, with naturally occurring empty capsids (80S, H antigen) and 14S precursor subunits, but not with heat-treated virion [80S, H(heated) antigen]. In contrast, Blondel et al. (1983) and Thorpe et al. (1982) described neutralizing monoclonal antibodies reactive with both N and H(heated) antigens. In this study, we isolated three hybridomas secreting monoclonal antibodies (2A 12, 4C4 and 6H8) which react with N and H(heated) antigenic particles of poliovirus, and confirmed that one of the antibodies, 4C4, had neutralizing activity.In another approach to investigate the antigenic properties of poliovirus, Urasawa et al. (1974, 1976) employed three kinds of equine serum inhibitor-resistant mutants of poliovirus Mahoney strain (M-HN31, M-HN11 and M-H33) which were obtained by propagating the strain in the presence of equine inhibitory sera (HN31, HNll and H33 respectively) with different neutralization specificities. By absorbing anti-Mahoney serum with the three inhibitor-resistant mutants, three kinds of neutralizing antibodies (HN31, HNll and H33 antibodies) recognizing different antigenic determinants of poliovirus were prepared (Urasawa et at., t974, 1976). In neutralization tests (Urasawa et al., 1979) and electron microscopy using the monospecific antibodies thus obtained~ it was found that one antibody (HN31 antibody...