Antigenic Analyses of Influenza Viruses by Complement Fixation

Abstract

Summary Antisera to the soluble complement-fixing S antigens of influenza viruses were readily produced in guinea pigs. Following intranasal instillation of active virus many animals developed antibodies to the virus V antigen, but the anti-S responses were variable as to incidence and height of titers obtained, depending to some extent upon the strain employed. The anti-S as well as anti-V formation was strikingly improved if the intranasal inoculation was followed in 2 wk or later by intra-abdominal injection of S antigen derived from the homologous strain. Anti-S titers of 1:64 or greater were thus obtained in more than 75% of the animals. As few as 100 infectious doses for chick embryos induced direct antibody formation in some, and conditioned additional animals to respond to the booster injection of S. Since more than a million times the amount of inactivated virus failed to yield either type of responses it may be assumed that guinea pigs sustain inapparent influenza infections. Intra-abdominal inoculation of active virus also led to anti-S responses, either directly or following secondary injections of S, but less frequently and to lower titers than in the intranasal series. In confirmation of previous reports no antigenic differences could be detected among S antigens derived from homotypic strains. In contrast, antibodies to the V antigens exhibited a considerable degree of strain-specificity, often to the extent that no cross-reactions could be discerned. The booster effects observed, both with respect to anti-S and anti-V, could not be ascribed to residual concentrations of hemagglutinating virus particles in the S preparations. It appears (a) that S, although shown to be incapable per se of inducting primary anti-S formation, even after numerous injections, provides an effective secondary antigenic stimulus in animals conditioned by previous infection; and (b) that the booster inocula contained some nondetectable V components, which were not linked to hemagglutinating particles. Sera containing anti-V in addition to anti-S can safely be used for analysis of S antigens if the test preparations are (a) rendered free of V by appropriate procedures; or (b) derived from strains with antigenically distinct V components. Safer yet, anti-V responses can be avoided in guinea pigs altogether, if strains with unrelated V antigens are employed for the primary infection and the preparation of the booster S antigen, respectively. By use of a small intranasal dose and by delay of the booster injection, high-titered anti-S sera were obtained, devoid of anti-V.