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全悬浮培养LMH细胞制备FAdV-4灭活疫苗的研究
秦红刚,韩兴,秦伟,李晶梅,熊亮,余继钦,谢红玲,漆世华,李凌峰,石宝兰
0
(国药集团动物保健股份有限公司)
摘要:
通过对LMH细胞进行驯化,获得一株可全悬浮培养的LMH细胞(命名为LMH-S),此细胞可适应无血清、高密度悬浮培养,以初始细胞密度0.5×106/ml~1×106/ml接种,细胞密度最高达6×106/ml。研究确定生物反应器悬浮培养LMH-S细胞制备FAdV-4抗原的最优参数为:接毒时细胞密度2×106个/ml~3×106个/ml、接毒量0.1 MOI~1 MOI、接毒后48 h~72 h收毒,所获抗原病毒含量不低于109.25 TCID50/ml。试验鸡分别免疫0.02 ml、0.05 ml、0.1 ml、0.3 ml疫苗,攻毒保护率分别为9/10、10/10、10/10、10/10,说明疫苗有效性良好。结果表明,本试验成功建立了LMH全悬浮培养细胞株,确定了FAdV-4抗原的生物反应器悬浮培养工艺,制备的FAdV-4灭活疫苗有效性良好,为高效FAdV-4灭活疫苗的研制奠定了基础。
关键词:  LMH细胞  悬浮培养  FAdV-4
DOI:
投稿时间:2023-05-04修订日期:2023-08-18
基金项目:
Preparation of FAdV-4 inactivated vaccine fromLMH cells in suspension culture
(Sinopharm Animal Health Co.LTD.)
Abstract:
we successful adaptation of adherent LMH cells to growth in suspension culture,named as LMH-S,which can cultured in serum-free miedium and growth to a maximum viable cell density of about 6×106/ml,when inoculation with the initial cell density of 0.5×106 cells /ml~1×106 cells /ml. wo also study the culturing procedure of FAdV-4 using this suspended LMH cells,which displayed that the best result could get when using the initial seeding density of 2×106 cells /ml~3×106 cells /ml, infected with 0.1 MOI~1 MOI, and harvested between post-infection 48 h~72 h.The titre of FAdV-4 virus in suspension culture of bioreaction was not less than 109.25 TCID50/ml.Immunized SPF chickens with this inactivated vaccine,The immune protection rate was 9/10、10/10、10/10 and 10/10 at the immune dose of 0.02 ml、0.05 ml、0.1ml and 0.3ml per chick,and the good immune protection effect was achieved. The results showed that wo successfully established the LMH cells which can cultured in suspension, and optimized the culture parameters of FAdV-4 use this LMH cells,wo also proved that the immune effection of FAdV-4 inactivated vaccine prepared by suspension culture technology was good, which provided a scientific basis for the large-scale production of vaccine.
Key words:  LMH  cells, Suspension  culture, FAdV-4

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