Takehara K, Hashimoto H, Ri T, Mori T, Yoshimura M
Laboratory of Poultry Diseases, School of Veterinary Medicine and Animal Sciences, Kitasato University, Aomori, Japan.
Virus Res. 1992 Nov;26(2):167-75. doi: 10.1016/0168-1702(92)90155-3.
With measles virus cDNA of the avirulent vaccine strain AIK-C, two cDNAs of H or F genes were amplified by the polymerase chain reaction. The amplified cDNAs were inserted respectively to the baculovirus transfer vector pAcYM1 derived from the nuclear polyhedrosis virus of Autographa californica (AcNPV). After co-transfection of the transfer vectors with AcNPV DNA to Spodoptera frugiperda cells, recombinant baculoviruses were screened by plaque assay, and the viruses containing H-cDNA or F-cDNA were named H-AIK or F-AIK, respectively. By Western blot analyses, the band around 80 kDa and some smaller bands were appeared in the H-AIK infected S. frugiperda cells, and the band around 40 kDa was detected in the F-AIK infected cells. Immunofluorescence studies on unfixed S. frugiperda cells infected with H- or F-AIK recombinants showed that both antigens were transported to the cell surface. When green monkey red blood cells were added to the recombinant infected cells, H-AIK infected cells showed haemadsorption, and cells infected with F-AIK lysed the red blood cells. The recombinant proteins elicited the neutralizing antibodies against measles virus.
利用无毒疫苗株AIK-C的麻疹病毒cDNA,通过聚合酶链反应扩增出H或F基因的两个cDNA。将扩增得到的cDNA分别插入到源自苜蓿银纹夜蛾核型多角体病毒(AcNPV)的杆状病毒转移载体pAcYM1中。将转移载体与AcNPV DNA共转染草地贪夜蛾细胞后,通过噬斑测定筛选重组杆状病毒,含有H-cDNA或F-cDNA的病毒分别命名为H-AIK或F-AIK。通过蛋白质免疫印迹分析,在感染H-AIK的草地贪夜蛾细胞中出现了约80 kDa的条带和一些较小的条带,在感染F-AIK的细胞中检测到了约40 kDa的条带。对感染H-或F-AIK重组体的未固定草地贪夜蛾细胞进行免疫荧光研究表明,两种抗原均转运至细胞表面。当将绿猴红细胞添加到重组感染细胞中时,感染H-AIK的细胞出现血细胞吸附现象,而感染F-AIK的细胞则使红细胞裂解。重组蛋白诱导产生了针对麻疹病毒的中和抗体。