Influenza A virus neuraminidase (NA) is drawing attention as a target for vaccine development. In this study, we propose kinetic MUNANA assay as a tool to identify monoclonal antibodies (mAbs) that specifically target functional epitopes on NA. By analyzing changes in the parameters of the Michaelis-Menten curve (Km and Vmax), we revealed distinct mechanisms of Ab-mediated inhibition. Additionally, we developed a web-based application facilitating efficient processing of the assay results and enabling statistical inference. We employed the kinetic MUNANA assay to test newly developed mAbs targeting NA of the widely used PR8 H1N1 strain. Among these, mAbs with strong effect on NA enzymatic parameters were more likely to select for escape mutants that had a substantial impact on the overall enzymatic activity of the virus. In summary, when combined with ELLA, kinetic MUNANA is a rapid method to profile the putative binding site and the effect of NA-specific mAbs.