Comparison of hemagglutination inhibition, single radial hemolysis, virus neutralization assays, and ELISA to detect antibody levels against seasonal influenza viruses

Background: The immunological response to influenza vaccine and/or natural infection is evaluated by serological techniques, the most common being hemagglutination inhibition (HI), single radial hemolysis (SRH), and virus neutralization assays, which is commonly used in a micro-neutralization (MN) format. ELISA is not officially required; however, this assay is able to measure different class-specific antibodies. The four assays identify different sets or subsets of antibodies. Objectives: The aim of this study was to establish the correlation among four serological assays using four seasonal influenza strains. Methods: The HI, SRH, MN assays, and ELISA were performed on four seasonal influenza strains. Results: A strong positive correlation was found between HI and MN and between SRH and MN assays for influenza A strains. The B strains also showed good correlations among the three assays. A positive correlation was also found between ELISA and the “classical” assays for all strains. Concerning the correlates of protection, as defined by HI ≥ 40 and SRH ≥ 25 mm2, good agreement was observed for the influenza A strains. By contrast, the agreement for the B strains was very low. Conclusions: There is a positive strong correlation among the four serological assays for both A and B strains, especially for the HI and MN assays. There is good agreement on correlates of protection between HI and SRH assays for the A strains, but very low agreement for the B strains, suggesting higher sensitivity of SRH than HI assay in detecting antibodies against the influenza B viruses.