Antigenic Analysis of Influenza Viruses by Complement Fixation

Abstract

Summary With the development of methods for the effective separation of the strain-specific virus (V) antigen from the broader reacting type-specific soluble (S) antigen and the production of sera containing antibodies to either one or the other of these antigens, it has become possible to demonstrate strain-specific components of influenza viruses by complement fixation. Data are presented on the application of strain-specific anti-V sera, free of anti-S, prepared against a variety of strains, to the identification and strain analysis of new isolates. The procedures used for definitive diagnosis depended upon whether the isolate was a member of a recognized family of strains, as A prime, or a totally new variant as represented by the Asian strains. Thus, three influenza viruses obtained from outbreaks which occurred in February 1957, on a preliminary screening test, were found to fix complement in the presence of several A prime antisera, but not with antisera against swine 15 or five prototype A strains. On titrating the cross-reacting antisera versus the new isolates, the results indicated that all three viruses were closely related to the Nederland 36 strain isolated in 1956. They were distinguishable from one another and from Nederland 36, however, by the spectrum and level of the cross-reactions with other A prime strains. Assay of antisera prepared against the new isolates further clarified their antigenic composition. When the Asian variant appeared, the identification of isolates could be restricted to the screening test since no cross-reactions were readily detectable with antisera against any of the earlier strains. Positive complement fixation occurred solely with the A/Japan/305 antiserum. Identification was carried out by testing aliquots of infected allantoic fluid, which contained at least 16 HA units, the equivalent of one V antigen unit, against a battery of anti-V sera diluted optimally in respect to their homologous reactions. In this fashion, early passage material containing as little as 40 HA units/ml could be identified, if 0.5 ml were used for testing. Thus, of 51 viruses obtained from throat washings 80% were diagnosed on first passage. The implications of the use of the technique are discussed.