Background <p>Hemagglutinin (HA), one of the main surface glycoproteins of the influenza virus, is essential for viral entry into host cells. Antibodies targeting HA can neutralize the virus, offering protection against infection. HA-specific antibody titers are therefore key markers of immune protection and vaccine efficacy. However, data on humoral immune responses to influenza A(H3N2) and influenza A(H1N1)pdm09 and viruses in India remain limited.</p> Methodology <p>Laboratory-confirmed influenza specimens from 2016 to 2017 were analyzed in this retrospective longitudinal study. A systematic random sampling method was used to obtain 160 paired serum samples, distributed among four distinct age groups. Hemagglutination inhibition (HAI) assays were performed to assess the antibody responses against influenza A(H3N2) and influenza A(H1N1)pdm09 viruses following natural infection. Additionally, preexisting antibodies and antibodies to heterologous influenza strains were evaluated.</p> Results <p>Following natural infection, seroconversion rates were 77.1% for individuals infected with influenza A(H3N2) and 84.9% for those infected with influenza A(H1N1). Preexisting antibodies were detected in 40.8% of the acute-phase influenza A(H3N2) samples and 17% of the acute-phase influenza A(H1N1)pdm09 samples. Antibodies to heterologous strains were also detected in both groups, particularly between influenza A(H1N1)pdm09 and influenza A(H3N2), as well as with influenza B lineages.</p> Conclusion <p>The results of this study show that most of the study population had evidence of immunity against both influenza A(H3N2) and influenza A(H1N1) virus strains. Evidence of antibodies to other influenza strains was also observed in a few samples, thus emphasizing the need for continuous monitoring and tailored vaccination strategies in India.</p>