摘要
为提高口蹄疫抗原的病毒含量,减少口蹄疫疫苗副反应,提升生产效率,降低生产成本,对口蹄疫病毒无血清悬浮培养生产工艺进行了研究。将BHK-21细胞在生物反应器中进行无血清悬浮培养,设计了最佳培养pH、接种细胞密度、培养温度、溶氧含量、接毒量、收获时间、成品免疫效果的试验方法,确定了生产工艺。结果表明,采用无血清悬浮培养工艺,按照接种细胞密度1.0×10^(6)/mL,培养pH为7.00~7.20,培养温度36.5~37.0℃,溶氧含量40%~50%进行BHK-21细胞培养,培养72h,细胞密度大于7×10^(6)/mL,沉降换液后,加入含2%口蹄疫种毒的病毒维持液进行培养后收获病毒液,口蹄疫抗原的病毒146S含量能达到18μg/mL以上,比传统低血清悬浮生产工艺提升了2倍以上,且制备的成品安全有效,免疫抗体阳性率为100%。本研究成功建立了适合口蹄疫病毒无血清悬浮培养的生产工艺,获得了更高效价的口蹄疫病毒,同时验证了无血清培养抗原的免疫效果,为高效价口蹄疫疫苗的生产和研究提供了新思路。
To improve the virus content of foot-and-mouth disease(FMD)antigen,reduce vaccine adverse reactions,enhance production efficiency,and reduce manufacturing costs.we conducted research on serum-free suspension culture production technology for FMD virus.BHK-21 cells were cultured in serum-free suspension with-in a bioreactor.Experimental protocols were designed to optimize culture parameters including pH,initial cell density,temperature,dissolved oxygen(DO),virus inoculation quantity,harvest timing,and final product immunogenicity.The optimal production process was established through systematic investigation.Results demonstrated that using serum-free suspension culture with optimized parametes(initial cell density 1.0×10^(6)/mL,pH 7.00~7.20,temperature 36.5~37.0℃,D0 40%~50%),cell density exceeded 7×10^(6)/mL after 72 hours of cultivation.Following sedimentation and medium exchange,infection with 2%FMD seed virus in maintenance medium yielded viral harvest.The resulting FMD antigen achieved 146S content exceeding 18μg/mL,representing a more than 2-fold increase compared to traditional low-serum suspension production.The prepared product is safe and effective with an immunological antibody positivity rate of 100%.This study successfully established an optimized serum-free suspension culture process for FMD virus production,obtained higher-titer viral antigen,and validated the immunogenic efficacy of serum-free cultured antigen,providing novel insights for high-titer FMD vaccine productiong and research.
作者
施程洪
李彦荣
陆礼琼
赵路路
杨建强
曾刚春
成建文
王继华
刘术新
SHI Cheng-hong;LI Yan-rong;LU Li-qiong;ZHAO Lu-lu;YANG Jian-qiang;ZENG Gan-cun;CHENG Jian-wen;WANG Ji-hua;LIU Shu-xin(Zhongpu Biopharmaceutical Co.,Ltd.,Baoshan,Yunnan 678000,China)
出处
《中国兽药杂志》
2025年第8期15-24,共10页
Chinese Journal of Veterinary Drug
基金
中普生物制药有限公司2024年科技攻关项目(ZPYF-2024-007)。
关键词
口蹄疫病毒
146S
无血清
悬浮培养
Foot-and-mouth disease virus
146S
serum-free
suspension culture
作者简介
施程洪,高级兽医师,硕士,从事动物疫苗生产管理及产品工艺研发工作;通讯作者:王继华。E-mail:bsymcwjh@sina.com。
