A study on the standard of influenza neuraminidase inhibition assay

In present study, standard method and standard operation practice for measuring the activities of influenza neuraminidase and its inhibitors have been established.  The accuracy and stability of the method has been evaluated.  Standard operation is as following: 10 μL sample, 30 μL neuraminidase and 60 μL substrate are added to one well of a 96-well plate, and then incubated at 37 ℃ for 1 h.  The reaction was stopped with NaOH before fluorescence intensity determination.  One unit of neuraminidase is defined as the amount of enzyme that produces 1 nmol 4-MU in 1 h under above conditions.  The inhibition accuracy is indicated by an uncertainty measurement of 6.51×10⁻², and its stability was reaffirmed by determination of oseltamivir acid.  In this study, systematic assessment of neuraminidase inhibitory assay not only provided theoretical basis of its application in drug discovery, but also made preliminary attempt to use uncertainty measurement as a parameter in biological measurement.