Radford A D, Willoughby K, Dawson S, McCracken C, Gaskell R M
Department of Veterinary Pathology, University of Liverpool, Veterinary Teaching Hospital, Leahurst, Neston CH64 7TE, United Kingdom.
J Virol. 1999 Oct;73(10):8496-502. doi: 10.1128/JVI.73.10.8496-8502.1999.
In order to map linear B-cell (LBC) epitopes in the major capsid protein of feline calicivirus (FCV), an expression library containing random, short (100- to 200-bp) fragments of the FCV F9 capsid gene was constructed. Analysis of this library showed it to be representative of the region of the capsid gene that encodes the mature capsid protein. The library was screened by using polyclonal antisera from a cat that had been challenged experimentally with F9 to identify immunoreactive clones containing LBC epitopes. Twenty-six clones that reacted positively to feline antisera in immunoblots were identified. FCV-derived sequence from these clones mapped to a region of the capsid that spanned 126 amino acids and included variable regions C and E. An overlapping set of biotinylated peptides corresponding to this region was used to further map LBC epitopes by using F9 antisera. Four principal regions of reactivity were identified. Two fell within the hypervariable region at the 5' end of region E (amino acids [aa] 445 to 451 [antigenic site (ags) 2] and aa 451 to 457 [ags 3]). However, the other two were in conserved regions (aa 415 to 421 [ags 1; region D] and aa 475 to 479 [ags 4; central region E]). The reactivity of the peptide set with antisera from 11 other cats infected with a range of FCV isolates was also determined. Ten of 11 antisera reacted to conserved ags 4, suggesting that this region may be useful for future recombinant vaccine design.
为了绘制猫杯状病毒(FCV)主要衣壳蛋白中的线性B细胞(LBC)表位图谱,构建了一个表达文库,该文库包含FCV F9衣壳基因的随机短片段(100至200 bp)。对该文库的分析表明,它代表了衣壳基因中编码成熟衣壳蛋白的区域。通过使用来自经F9实验性攻击的猫的多克隆抗血清筛选该文库,以鉴定含有LBC表位的免疫反应性克隆。在免疫印迹中对猫抗血清呈阳性反应的26个克隆被鉴定出来。来自这些克隆的FCV衍生序列映射到衣壳的一个区域,该区域跨度为126个氨基酸,包括可变区C和E。使用与该区域对应的一组重叠的生物素化肽,通过使用F9抗血清进一步绘制LBC表位图谱。确定了四个主要反应区域。两个位于区域E 5'端的高变区内(氨基酸[aa]445至451[抗原位点(ags)2]和aa 451至457[ags 3])。然而,另外两个位于保守区域(aa 415至421[ags 1;区域D]和aa 475至479[ags 4;中央区域E])。还测定了该肽组与来自感染一系列FCV分离株的其他11只猫的抗血清的反应性。11份抗血清中有10份与保守的ags 4发生反应,这表明该区域可能对未来的重组疫苗设计有用。