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Growing interest in universal influenza vaccines and novel administration routes has led to the development of alternative serological assays that are able to detect antibodies against conserved epitopes. We present a competitive ELISA method that is able to accurately determine the ratio of serum immunoglobulin G directed against the different domains of the hemagglutinin, the head and the stalk. Human serum samples were treated with two variants of the hemagglutinin protein from the A/California/7/2009 influenza virus. The signals detected were assigned to different groups of antibodies and presented as a ratio between head and stalk domains. A subset of selected sera was also tested by hemagglutination inhibition, single radial hemolysis, microneutralization, and enzyme-linked lectin assays. Pre-vaccination samples from adults showed a quite high presence of anti-stalk antibodies, and the results were substantially in line with those of the classical serological assays. By contrast, pre-vaccination samples from children did not present anti-stalk antibodies, and the majority of the anti-hemagglutinin antibodies that were detected after vaccination were directed against the head domain. The presented approach, when supported by further assays, can be used to assess the presence of specific anti-stalk antibodies and the potential boost of broadly protective antibodies, especially in the case of novel universal influenza vaccine approaches.

Influenza Anti-Stalk Antibodies: Development of a New Method for the Evaluation of the Immune Responses to Universal Vaccine