基于抗原蛋白Fiber-2及细胞因子佐剂的Ⅰ群4型禽腺病毒亚单位疫苗的开发

为了开发一种抗I群4型禽腺病毒(FAd V-4)的亚单位疫苗,选择I群禽腺病毒血清4型流行毒株衣壳蛋白Fiber-2作为保护性抗原蛋白。利用昆虫细胞-杆状病毒系统生产Fiber-2蛋白,通过Westernblot检测重组蛋白的表达情况,利用双向琼脂扩散法检测重组Fiber-2的免疫原性。为了增强亚单位疫苗的免疫保护效果,添加鸡源白细胞介素2(chIL-2)和鸡源γ-干扰素(ch IFN-γ)作为免疫佐剂,与重组Fiber-2单独或混合使用,制备多种形式的亚单位疫苗,并进行动物攻毒保护试验。重组蛋白的Western-blot鉴定结果显示,63 ku处出现Fiber-2蛋白目的条带;双向琼脂扩散试验结果显示,重组蛋白效价达到1∶128,表现出良好的反应原性。动物攻毒保护试验结果显示,单独使用Fiber-2免疫SPF鸡,在50 μg/只的剂量下,可为SPF鸡提供100%的保护效果,以chIL-2和chIFN-γ为免疫佐剂,与重组Fiber-2联合免疫SPF鸡,可显著增强Fiber-2在低剂量(10 μg/只)下的免疫保护效果,上述结果表明重组Fiber-2联合chIL-2和chIFN-γ细胞因子佐剂所制备的亚单位疫苗在抗I群4型禽腺病毒感染领域具有良好的应用前景。

Development of group Ⅰ serotype 4 fowl adenovirus subunit vaccine based on antigen protein Fiber-2 and cytokine adjuvants

To develop a subunit vaccine against group Ⅰ serotype 4 fowl adenovirus(FAdV-4),Fiber-2 was selected as the protective antigen.The recombinant Fiber-2 was expressed in insect cells using the baculovirus expression vector system.The expression of recombinant Fiber-2 was detected by Westernblot.The immunogenicity of recombinant Fiber-2 was detected by two-way agar diffusion method.Then the recombinant chicken IL-2(chIL-2) and recombinant chicken IFN-γ(chIFN-γ) were added as immunological adjuvant producing various forms of subunit vaccines.The immune efficacies of these different subunit vaccines were evaluated by animal experiment.Western-blot showed there was an obvious band at proximately 63 ku as we expected.The results of two-way agar diffusion test showed the recombinant protein titer reached 1 ∶ 128,showing well reactogenicity.The animal experiment result showed these subunit vaccines had an effective protection for SPF chickens.When injecting the recombinant Fiber-2 alone at 50 μg each bird could provide 100% protection for chickens.When adding chIL-2 and ch IFN-γ as adjuvant could significantly enhance the protective effect of the recombinant Fiber-2 at low doses(10 μg each bird).These indicate that recombinant Fiber-2 protein combine with chIL-2 and ch IFN-γ cytokine subunit vaccines have perfect application prospects against group Ⅰ serotype 4 fowl adenovirus infection.