Sia cleavage preference and susceptibility to oseltamivir inhibition. (A) The cleaving preference of IAV NA was tested by incubation of IAV with the fluorescent reporter, 4-methyl-umbelliferyl (4MU), linked to Neu5Ac, Neu5Gc, or 2-keto-3-deoxynononic acid (Kdn) as substrate. All virus strains cleaved Neu5Ac and Neu5Gc but not Kdn. Background from spontaneous degradation of the 4MU-Sia compounds was subtracted from the results. (B) Virus susceptibility to oseltamivir inhibition was tested by incubation of IAV with 2.5 nmol 4MU-Neu5Ac in the presence of 0, 1 or 17.5 μM oseltamivir in triplicates. Certain strains were only partially inhibited by 1 μM oseltamivir, however, all of the strains were inhibited by 17.5 μM oseltamivir. Substrate cleavage was quantified by measuring fluorescence from the released 4MU reporter compound. *P<0.05, **P<0.01 (two-tailed T-Test, n=3).