The specificity of RWJ-270201 as an influenza virus neuraminidase inhibitor was investigated by determining its inhibitory activity against neuraminidases from a variety of sources in an in vitro enzymatic assay. The IC50 of RWJ-270201 against neuraminidases derived from other sources, including mammalian (rat liver), bacterial (V. cholerae and C. perfringens), and other viral (parainfluenza virus and Newcastle disease virus) neuraminidases was >300 μM, which is at least 4 orders of magnitude less potent than that for influenza virus neuraminidase. This indicates that RWJ-270201 is a highly selective inhibitor for influenza A and B neuraminidases.