Antigenic Analysis of Influenza Viruses by Complement Fixation
Authors: Lief Florence S. , Henle Werner 
Published: January 2023

Summary
The age distribution of complement-fixing antibodies to specific virus (V) antigens of prototype strains of influenza Type A virus isolated between 1931 and 1957 was determined using acute and convalescent stage sera of patients suffering from A1 or A2 influenza, and pre- and postvaccination sera of persons injected subcutaneously with monovalent A2 virus vaccine.
On the basis of incidence of positive reactions at serum dilutions of 1:4 or greater, many individuals possessed or developed antibodies to V antigens of strains to which they could not have been exposed during their lifetime; they were born either long after a given strain was circulating or the strain became prevalent only at some time after certain of the sera were collected. For example, many young children revealed or formed antibodies to swine influenza and A subtype strains, and a number of individuals above 10 years of age possessed, prior to the dissemination of the A2 virus, antibodies to this variant in their acute stage or prevaccination sera or produced them in response to an A1 infection.
On the basis of geometric mean serum titers in the acute stage and prevaccination sera, the age distribution of antibodies to the prototype viruses appeared to be more clearly related to the era of prevalence of the strains although there was considerable overlapping. According to this kind of evaluation, children seemed to possess antibodies only to A1 strains; young adults revealed antibodies to swine influenza A and A1 strains in nearly equal concentrations; persons over 30 years of age showed predominantly antibodies to swine influenza but antibodies to all other strains were also present to a slightly lesser extent, and only individuals above 50 years old harbored enough anti-A2V in their sera to be reflected in the geometric mean level. There was some indication that maximal mean titers occurred in those segments of the populations which were in their second decade of life when the particular agents were circulating.
The differences in age distribution largely disappeared in convalescence, particularly above the age of 7 years. Marked levels of antibodies were formed to all the V antigens tested whether patients were 7–10 years or 41–50 years of age. In the very young, however, the responses were often, but by no means always, restricted to a few antigens if not solely to the infecting virus. In older patients the responses, likewise, were on occasion limited in breadth. Vaccination rarely induced formation of antibodies to heterologous strains unless those antibodies were detectable in the serum taken prior to immunization. Antibodies to swine influenza virus were the exception. Heterologous antibodies present in prevaccination sera were readily boosted by injection of the A2 virus vaccine.
The significance of these various findings has been discussed in relation to the epidemiologic history and the antigenic composition of influenza viruses. To explain the data it is postulated that in addition to the dominant and minor antigens, detectable by in vitro tests, each homotypic variant must also contain antigens representative of many if not all past and even future strains. These hidden antigens are most likely present in trace quantities and may stimulate homologous antibody formation under the following conditions: a) they may readily recall antibodies in individuals who had been previously exposed to strains of influenza virus in which they were dominant; b) they may condition individuals on repeated infections to ultimately respond with detectable levels of antibodies; and c) they may even reach, in severe first influenza infections, an antigenic mass sufficient to stimulate a primary antibody response along with the expected reaction to the dominant and minor antigens of the infecting strain.