基于Box-Behnken实验设计与响应面法优化禽流感疫苗生产工艺?

Optimization of avian influenza vaccine culturing by Box-Behnken design and response surface methodology

为优化基于MDCK细胞无血清悬浮培养生产禽流感病毒的工艺以提高禽流感病毒血凝素(HA)滴度,采用部分析因实验设计考察了病毒生产工艺中接毒时细胞密度、TPCK-胰酶浓度、培养温度、感染复数和病毒收获时间对病毒HA滴度的影响,确定了接毒时细胞密度、TPCK-胰酶浓度、和病毒收获时间作为影响病毒HA滴度的显著因子,并进一步采用3因素3水平的Box-Behnken实验设计和响应面法,对3个显著因子作用条件进行了优化,获得了最优的工艺条件。结果显示:当接毒时细胞密度为7.0×10~6 cells×m L^(-1)、TPCK-胰酶浓度为17 mg×L^(-1)、病毒收获时间为60 h时,病毒HA滴度最高可达10.85 log_2 HAU/25μL。以上研究结果为禽流感疫苗高效大规模工业化生产提供了有力的数据支持。

In order to optimize production processes of avian influenza vaccine based on MDCK cell serum-free suspension culture, and improve avian influenza virus Hemagglutinin (HA) titer, Effect of cell density at infection, TPCK-trypsin concentration, Temperature post infection, multiplicity of infection and virus harvesting time were investigated by fractional factorial design. Cell density at infection, TPCK-trypsin concentration, and virus harvesting time were determined as significant factors affecting the virus HA titer, and Box-Behnken experiment design with three factors and three levels together with response surface methodology were further applied. Virus HA titer of 10.85 log 2 HAU/25 μL can be obtained under the optimum condition of cell density at infection = 7.0 × 106 cells·mL-1, TPCK-trypsin concentration = 17 mg·L-1 and virus harvesting time = 60 h. These results provide fundamental data for efficient and large-scale industrial production of avian influenza vaccine.