# Influenza neuraminidase active site proximity assay for rapid profiling of inhibitory antibodies and antigenic drift

## TL;DR

A new assay called NASPA helps quickly measure antibodies against influenza neuraminidase, aiding vaccine strain selection and tracking antigenic changes.

## Contribution

NASPA is a novel, rapid, and cost-effective solution-based assay for profiling inhibitory antibodies against influenza neuraminidase.

## Key findings

- NASPA titers correlate strongly with ELLA titers and are not affected by anti-HA antibodies.
- NASPA detected incremental antigenic changes in recent B Victoria lineage vaccine strains.
- Steric and active site-binding NAI antibodies against circulating NAs are common in adult human sera.

## Abstract

Efficient approaches that can help to select vaccine strains for the influenza virus neuraminidase (NA) antigen are currently needed to advance the development of vaccines containing NA. Here, we present a rapid and cost-effective solution-based NA active site proximity assay (NASPA) for measuring NA activity inhibitory (NAI) antibodies. This simplified assay uses large “bulky” NA active site-binding inhibitors to replace the sialylated glycoprotein substrates in common NA enzyme-linked lectin assay (ELLA) approaches. Our results with ferret antisera and monoclonal antibodies against vaccine strain NAs show a strong correlation between NASPA and ELLA titers, and that NASPA titers are not influenced by anti-HA antibodies. Consequently, NASPA can be used with influenza A or B strains and with the latter it revealed incremental antigenic changes in the NAs from recent B Victoria lineage vaccine strains. By coupling NASPA with a simple activity assay, we also found that steric and active site-binding NAI antibodies against circulating NAs are common in adult human sera. Finally, we demonstrate that NASPA can be modified by incorporating novel NA substrate-analog-based inhibitors. Together, these results suggest that NASPA can aid the development of vaccines containing NA by helping to select suitable vaccine strains and profile anti-NA antibody responses.