RESUMO
To simplify the preparation of neuraminidase in screening influenza neuraminidase inhibitors,the neuraminidase gene of H5N1 influenza A virus was optimized for high expression in mammalian cells and cloned into pcDNA4/TO vector. The recombinant plasmids were transfected into T-REx 293 cells to establish stable cell lines,in which the expression of neuraminidase was induced by tetracycline. Unlike from virions,the preparation of neuraminidase became conveniently and safely from these stable cell lines,which would facilitate developing high throughput assay to screen neuraminidase inhibitors. More than 3000 natural extracts and herbal components were screened in the study. Baicalin and baicalein were found to inhibit oseltamivir-sensitive and oseltamivir-resistant neuraminidase at similar level,furthermore,their anti-influenza activity was confirmed by plaque assay and virus inhibition assay.