PMC

Purified E59/EV71 was inactivated by 0.025% (v/v) formalin at different temperature and the residual of virus infectivity was detected by plaque assay performed triplicate at different time points.

Vero cells (2×10⁵ cell/ml) was cultured in 75T-flask and counted for viable cell number every 24 hours (0, 24, 48, 72, 96, 120 and 144 hours). The conditions for cell culture were described in the section.

(A) The T-flask was seeded with either 1.0∼1.5×10⁶ or 2.0∼2.5×10⁶ Vero cells, then after 2–3 days the cells were infected by the different ratio of E59/EV71 virus. The effects of different M.O.I were detected in the kinetic profile of virus produced from Vero cell grown in the VP-SFM medium. (B) The consistency of 3 Lots of EV71 virus production in the roller bottles. Virus titer was detected every day by TCID50 for 5 days.

Lane M is MW Markers; lane 1 is EV71 vaccine product produced from serum-containing medium and stored at 4°C for 26 months; lanes 2 and 3 are EV71 vaccine products derived from Lot #1 and 2 and stored at 4°C for 13 and 4 months, respectively. Monoclonal antibody used in the Panels B and C are MAB979 specific for VP2 and N16 specific for VP1, respectively.