MDCK and the stable shRNA expressing cell lines in 24-well plates were infected with viruses at moi of 0.005,0.5 (2 mg/mL trypsin was used in the infection process of H1N1). After incubation for 1 hr, the infected medium was removed and MEM without FBS was added. Cell supernatants were collected at different time points. The viral load was detected by hemagglutination (HA) and/or plaque assays as described previously [39] . Briefly, the HA assay was carried out in U-bottom 96 well plates. Serial 2-fold dilutions of virus samples were mixed with an equal volume of a 0.5% suspension of turkey erythrocytes (Lampire Biologic Laboratories, Pipersville, USA) and incubated at room temperature (RT) for 45 mins. Wells containing an adherent, homogeneous layer of erythrocytes were scored as positive. For plaque assay, serial 10-fold dilutions of virus sample were added into a monolayer of MDCK cells. After 1 hr incubation, the virus was removed and the cultures were overlaid with 1% semi solid agar-MEM. Three days after infection, plaques were visualized by staining of crystal violent.