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0.5×10^(6)/mL~1×10^(6)/mL接种,细胞密度最高达6×10^(6)/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2×10^(6)/mL~3×10^(6)/mL、接毒量0.1MOI~1MOI、接毒后48h~72h收毒,所获抗原病毒含量不低于10^(9.25)TCID_(50)/mL。试验鸡分别免疫0.02mL、0.05mL、0.1mL、0.3mL疫苗,攻毒保护率分别为9/10、10/10、10/10、10/10,表明疫苗有效性良好。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,验证了悬浮培养抗原的免疫原性,为高效FAdV-4灭活疫苗的研制奠定了基础。
The LMH cells was successful adapted to growth in suspension culture,named as LMH-S,which can cultured in serum-free medium and growth to a maximum viable cell density of about 6×10^(6)/mL,when inoculation with the initial cell density of 0.5×10%/mL~1×10^(6)/mL.The culturing procedure of FAdV-4 using these suspended LMH cells also studied,which displayed that the best result could get when using the initial seeding density of 2×10^(6)/mL~3×10^(6)/mL,infected with 0.1MOI~1MOI,and harvested between post-infection 48 h~72 h.The titer of FAdV-4 virus in suspension culture of bioreaction was not less than 10^(9.25)TCID_(50)/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 mL0.05 mL.0.1 mL and 0.3 mL per chick,the good immune protection effect was achieved.The results showed that the LMH cells which can cultured in suspension was successfully established,the culture parameters of FAdV-4 use this LMH cells also optimized,and immunogenicity of suspension culture antigen was also verified,which provided a scientific basis for the large-scale production of vaccine.
作者
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
QIN Hong-gang;HAN Xing;QIN Wei;LI Jing-mei;XIONG Liang;YU Ji-qin;XIE Hong-ling;QI Shi-hua;LI Ling-feng;SHI Bao-lan(Sinopharm Animal Health Corporation Ltd.,Wuhan 430075,China)
出处
《中国兽药杂志》
2023年第9期7-13,共7页
Chinese Journal of Veterinary Drug
