将中和性流行性感冒 (流感 )病毒基因工程抗体IV 2、IV 6的轻链和重链Fd段基因 ,分别克隆入全抗体表达载体 pAC L Fc ,构建成杆状病毒表达载体pAC L Fc Ⅳ 2和 pAC L Fc Ⅳ 6 ,转染昆虫Sf9细胞 ,利用杆状病毒 /昆虫细胞系统实现抗体...将中和性流行性感冒 (流感 )病毒基因工程抗体IV 2、IV 6的轻链和重链Fd段基因 ,分别克隆入全抗体表达载体 pAC L Fc ,构建成杆状病毒表达载体pAC L Fc Ⅳ 2和 pAC L Fc Ⅳ 6 ,转染昆虫Sf9细胞 ,利用杆状病毒 /昆虫细胞系统实现抗体的分泌型表达 ,表达产物进行亲和层析分离纯化。SDS PAGE电泳和Westernblot法证实有完整免疫球蛋白的表达 ,免疫印迹法证实它们能与流感病毒血凝素蛋白特异性结合。经间接竞争性抑制ELISA法测定 ,抗体与流感病毒抗原结合的解离常数KD 值分别为 2 5× 10 -9M和 3 0× 10 -9M。流感病毒基因工程全抗体经在昆虫细胞中的表达、纯化和抗体特性鉴定 ,获得了两株纯化的全抗体 ,可用于以后的动物模型呼吸道粘膜被动免疫抗感染的研究。展开更多
YN87448 virus strain was isolated from a fevered female patient (52 years old ) in Yunnan Province in 1986, and was identified as a member of Alphavirus using serological method. One primer was designed from common ha...YN87448 virus strain was isolated from a fevered female patient (52 years old ) in Yunnan Province in 1986, and was identified as a member of Alphavirus using serological method. One primer was designed from common hairpin conserved region of Alphavirus. Two fragments were amplified by single primer differential RT PCR, and they were cloned into pGEM T vector. Sequences were determined, and then analyzed by Software and GenBank. Results showed that the 1118pb long fragment was highly homologous to the sequence of Sindbis like virus S.A.A.R86. The homogeneity of the 1118bp fragment between YN87448 virus strain and S.A.A.R86 virus strain was 98%. Single primer differential RT PCR is a useful method with simple, economic, and practical value for Alphavirus identification.展开更多
文摘YN87448 virus strain was isolated from a fevered female patient (52 years old ) in Yunnan Province in 1986, and was identified as a member of Alphavirus using serological method. One primer was designed from common hairpin conserved region of Alphavirus. Two fragments were amplified by single primer differential RT PCR, and they were cloned into pGEM T vector. Sequences were determined, and then analyzed by Software and GenBank. Results showed that the 1118pb long fragment was highly homologous to the sequence of Sindbis like virus S.A.A.R86. The homogeneity of the 1118bp fragment between YN87448 virus strain and S.A.A.R86 virus strain was 98%. Single primer differential RT PCR is a useful method with simple, economic, and practical value for Alphavirus identification.
