Evaluation of the Efficiency of Polyclonal Antiserum Against the Conserved Motif of Influenza Virus Hemagglutinin in the Detection of Homologous and Heterologous Viruses

The H9N2 avian influenza is one of the most important viral diseases of poultry in Asian countries. Continuous mutations in the hemagglutinin (HA) protein as the main viral antigen make interpreting the HA-based serology test results difficult.

In this experimental study, the ability of polyclonal antibodies against the conserved motif of HA protein in detecting heterologous H9N2 isolates was evaluated.

Based on the bioinformatics data, a conserved motif of HA protein was selected and expressed in the pET-28a(+) vector. The recombinant peptide was mixed with an oil adjuvant and injected into specified pathogen free (SPF) chickens to produce the polyclonal antiserum. Similarly, antiserum from the inactivated virus was prepared. The efficiency of the prepared antisera was evaluated in the cross hemagglutination inhibition (HI) test using the homologous and heterologous H9N2 viruses.

The average titer of HI antibody against homologous and heterologous viruses using the polyclonal antiserum was equal to five. This titer was estimated to be seven for the homologous virus and much lower for heterologous viruses with inactivated virus antiserum. The difference was statistically significant.

According to the evaluations, this antiserum has a suitable efficiency in identifying different isolates of the H9N2 virus, and compared to the usual antiserum, it detects different antigens of a subtype more accurately. The resulting polyclonal antiserum does not contain non-specific inhibitors, which may cause problems interpreting serological findings. The results showed that polyclonal antiserum could identify H9N2 viruses isolated in different years by HI test; however, its validation requires testing with more viruses.