生物反应器培养轮状病毒基因重配株Ls条件的优化

Optimization of technological procedure on the reassortant rotaviruses strain Ls cultivation inbioreactor

目的轮状病毒基因重配株Ls(G3型)在生物反应器微载体培养Vero细胞条件的优化。方法采用3 L生物反应器微载体培养Vero细胞,观察Ls株在不同病毒感染复数(0.001、0.002、0.010、0.040 MOI)、不同温度(34.5℃和35.5℃)、不同病毒收获时间(24和48 h)对病毒增殖的影响。根据病毒滴度和收获量筛选出最适MOI、培养温度及病毒的收获时间。结果以0.002 MOI接种Vero细胞,温度为34.5℃培养病毒,滴度最高达7.50 lg CCID50/m L;48 h可连续收获4次病毒液,且收获总量及病毒滴度均高于24 h。结论通过对Ls株在生物反应器微载体Vero细胞培养条件的优化,获得的病毒液滴度高及连续培养多次收获量增加的有效方法,为进一步规模化培养奠定了基础。

Objective To optimize the key condition for culture of rotavirus reassortant strain Ls（ Type 4） by using vero cells adsorpted on microcarriers cultured in bioreactor.Methods The effect to multiplication of the virus was investigated, Vero cells cultured in 3 L bioreactor as cell matrix, then inoculated with Ls strain at MOIs of 0.001, 0.002, 0.010 and 0.040, in different temperatures（34.5℃or 35.5℃） and different harvest intervals（24 h or 48 h）, on which the optimal MOI, temperature and havest interval were decided.Results Under the condition of 0.002 MOI, 34.5 ℃, Ls was cul-tured well with a titer up to 7.50 lgCCID50/mL, and a continuous four harvests obtained in 48 h, and the total harvest and the virus titers were higher than cultured in 24 h.Conclusion An effective method for increasing rotavirus titer was devel-oped by optimizing the condition for culture of Ls on Vero cells in biorector,on which the foundation was developed in a large scale preparation of rotavirus vaccine, live.