Preparation of sera for subtyping of influenza A viruses by immunofluorescence

The conditions for preparation of type-specific and subtype-specific influenza A virus reagents to be used in the immunofluorescence technique have been evaluated. Type A-specific antibodies were prepared by passing an antivirion hyperimmune serum through an immunoadsorbent column containing antigens from disrupted virions of a different influenza A virus subtype. The type-specific antibodies were recovered from the immunoadsorbent by desorption with 3 M NaI. For subtype determination, antisera against the various hemagglutinins were used. Such sera could be prepared by removal of irrelevant influenza A virus antibodies from sera directed against purified virions and isolated peplomers, respectively. This was performed by passing the antisera through immunoadsorbent columns containing antigens from disrupted virions of appropriate strains. However, attempts to obtain a hemagglutin-specific antiserum from a serum directed against allantoic fluid virus suspension failed with this procedure. Antisera obtained after immunization with purified hemagglutinin were also elaborated. These sera were shown to be superior for subtyping of influenza A virus infections by immunofluorescence, but could not a priori be regarded as subtype-specific. The usefulness of subtype-specific sera has been demonstrated on clinical specimens for rapid virus diagnosis.