为研究由悬浮培养的 MDCK 细胞制备的 H9亚型 AIV 抗原所制备的灭活疫苗的免疫剂量，在生物反应器中悬浮培养 MDCK 细胞，接种 H9亚型禽流感病毒 WD98和 HN04种毒，收获抗原液，以2种抗原液分别制备2个毒株的单价油佐剂灭活疫苗。每种疫苗均按照0．3、0．2、0．1、0．05 mL／只（107．5 EID50／只～105．52 EID50／只）的剂量接种21日龄 SPF 鸡，免疫接种后3周采血测定 AIV 的 HI 抗体，采血后分别用与制苗株同源的 H9亚型 AIV 种毒静脉注射攻毒，测定疫苗保护率。2种疫苗不同剂量免疫接种后21 d 的 AIV HI 抗体水平达到3．8log2～5．6log2，对照组鸡 AIV HI 抗体均为0log2。攻毒试验证实，以105．52 EID50／只～107．5 EID50／只剂量进行免疫接种，免疫鸡只获得了80％以上的保护。按照兽药典 H9亚型禽流感疫苗攻毒保护率90％为合格的标准，悬浮培养 MDCK 细胞制备的 H9亚型禽流感疫苗的最小免疫接种剂量为106．82 EID50／只。%The immune dose was studied to the MDCK cell suspension cultured H9 subtype AIV inactivated vaccine.MDCK cells was suspensibly cultured in the bioreactor and inoculated with H9 subtype AIV of WD98 and HN04 strains.Antigen was harvested and with which the oil adjuvant inactivated vaccine was prepared.21 days-old SPF chickens were injected subcutaneously to the neck of the chickens with 0.3,0.2, 0.1,0.05 mL/per chicken (107.5 EID50/ per chicken-105.52 EID50/ per chicken)of the two H9 subtype AIV inactivated vaccine.3 weeks after immunization,blood samples were collected and chickens were challenged with the AIV which was same strain to the vaccine antigen by intravenous route.The blood samples were used for the HI titer test and the challenge chickens were used to evaluate the protection rate.The results showed that,21 days after immunization,the average HI titers of the vaccine groups were 3.8log2-5.6log2 of the two vaccines,and the AIV HI titer in blank control group of chicken was 0log2.The challenge test confirmed,when chickens immunized with 105.52 EID50-10 7.5 EID50/per chicken,the protection rate were 80%-100%.The result showed that when the chickens were immunized with 106.82 EID50 per chicken the protection rate reached 90%-100%.Accoding to veterinary pharmacopoeia standard of the challenge protec-tion rate of 90% of the H9 subtype avian influenza vaccine,the minimum immune dose of suspension cul-ture H9 subtype avian influenza vaccine was 106.82 EID50 per chicken.
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