微载体灌注培养Marc-145细胞制备猪繁殖与呼吸综合征疫苗

Preparation of PRRS Vaccine by Perfusion Culture of Marc-145 Cells on Microcarriers

关键词:  微载体  灌注培养  猪繁殖与呼吸综合征疫苗

Key words:  microcarrier  perfusion culture  PRRS vaccine

摘要:

对在生物反应器中用微载体连续灌注培养Marc-145细胞生产猪繁殖与呼吸综合征病毒的制备技术进行了研究。在14 L体积的生物反应器中，加入含10 g/L微载体的细胞培养基DMEM，接种Marc-145细胞至细胞浓度为1?105/mL，培养4 d后细胞可生长至5～7?106/mL，然后以感染复数(MOI)为0.01接种PRRSV PC株病毒，接毒后36 h开始收获，连续收获3 d左右，收获的病毒滴度范围在106.0～107.3TCID50/mL之间，将收获的病毒液加入适量的保护剂，经冷冻干燥制备成疫苗，无菌、支原体等项目的检验均合格，3批疫苗的免疫保护率均为5/5。实验表明，用生物反应器微载体灌注培养Marc-145细胞制备PRRS疫苗工艺可行。

Abstract:

A technique for preparing porcine reproductive and respiratory syndrome(PRRS) vaccine by perfusion culture of Marc-145 cells on microcarriers in bioreactors was established. A 14-liter bioreactor was filled with DMEM cell culture medium which contained 10 g/L of microcarriers, Marc-145 cells were inoculated with a final cell density of 1?105/mL. After 4 days of culture，cell density can reach 5?106～7?106/mL. Then PRRSV PC strain was inoculated with a MOI of 0.01. After inoculation for 36 hours, the cultured cells were harvested continuously for 3 days. The titer of virus in the culture medium was about 106.0 to 107.3TCID50/mL, which was then added with suitable amount of protective agent and freeze-dried to form vaccine. The aseptic, Mycoplasma and other tests were used to qualify. The immune protective rate of the three batches vaccine was 5/5, which demonstrated that the preparation technology of PRRS vaccine by perfusion culture of Marc-145 cells on microcarriers in bioreactors was viable.