Development Of BHK Cells Suspension Culture Process For The Production Of Foot-and-Mouth Vaccine

As one of the most infectious diseases, the outbreak of Foot-and-Mouth Disease will cause huge economic losses to animal husbandry and even the national economy. Safe and effective vaccine is one of the most important measures to prevent the disease. The use of traditional roller-bottle production process often brings some problems, such as complicated operation, difficulty in scale-up, and low production rate. In addition, the use of serum will inevitably lead to a rise in the cost of Foot-and-Mouth vaccine, enhancement of safety danger in the production process, and difficulty in controlling the quality of the vaccine. Therefore, there is an urgent need to develop a serum-free culture medium suitable for suspension cultures of BHK cells and a suitable virus production process.Firstly, a serum-free medium applicable for BHK cell growth was developed according to the mixture design and response surface design method. The serum-free medium can support cell growth to a maximum living cell density of (8.66±0.07)×106 cells/ml. Secondly, the suspension adaptation of BHK adherent cells was carried out using the optimized non serum medium, and the morphology, growth and metabolism regulation and production capacity of BHK suspension cells were investigated.10 days were needed to make BHK cells adapt suspension culture using the optimized serum-free culture medium. The BHK suspension cells after successful domestication could grow stably, and the average specific growth rate was about 0.90 d-1. The BHK suspension cells had good shape, uniform size, smooth and clear contour. Compared with the low serum commercial culture medium, the optimized serum-free culture medium gained 24.79% higher the maximum viable cell density of the cells and the stationary phase of the cells was prolonged by 2 days. Finally, the effect of inoculum density and liquid change inoculation on virus amplification in the cells was preliminarily studied. The results showed that the optimal initial inoculum density was 0.5×106 cells/ml. In addition, using the optimal initial inoculum density and liquid change inoculation, the virus titer of optimized serum-free medium and low serum commercial medium can reach 7.0 lgTCID50/0.1 ml and 7.12 lgTCID50/0.1 ml。By contrasting the virus titer and cell lesion rate using two kinds of culture media, we could find that the optimized serum-free medium was able to reach the same level both in virus titer and the cell lesions rate with the low serum commercial culture. The optimized serum-free medium can be used in the virus production.Based on the content of this article, we can develop a serum-free medium applicable for Foot-and-Mouth vaccine production, a kind of BHK suspension cells domesticated to grow in serum-free medium, preliminarily explore the feasibility of serum-free suspension FMD vaccine production process, which provide reference and data support for the industrial production of the Foot-and-Mouth vaccine.