中文摘要：

目的制备2型猪链球菌（S．suis2）表面抗原一（Sao）重组蛋白的多克隆抗体，鉴定其免疫学特性。方法通过PCR从2型猪链球菌中国强毒株05ZYH33基因组中扩增基因Sao，构建重组表达质粒pET28a—Sao，转化EcoliBL21，异丙基硫代半乳糖苷诱导表达目的蛋白，十二烷基磺酸钠一聚丙烯酰胺凝胶电泳鉴定表达产物，His亲和层析柱纯化重组蛋白，利用纯化后Sao蛋白制备多克隆抗体，间接酶联免疫吸附试验检测抗体效价，West—ernblot鉴定抗体特异性，评价Sao多克隆抗体的促全血杀菌作用。结果构建的重组质粒可以在宿主菌中高效表达，纯化后获得的重组蛋白纯度可达90％；间接酶联免疫吸附试验测定Sao多克隆抗体效价为1：102400；Westernblot结果显示，Sao多克隆抗体有较好的特异性；全血杀菌实验显示，05ZYH33在含有Sao多抗血清的全血中相对存活率下降85％。结论本研究制备的2型猪链球菌重组Sao蛋白多克隆抗体效价高，特异性好，可明显增强全血对细菌的杀伤作用。

英文摘要：

Objective To express the gene encoding surface antigen one （Sao） in highly virulent strains of Strepto- coccus suis serotype 2 （ S. suis 2） and to prepare the polyclonal antibodies against Sao to assess the immune characteristics of the anti-Sao polyclonal antibodies. Methods The target gene sao was amplified by PCR using the genomic template of 05ZYH33. Sao gene was subsequently inserted into pEASY-Tl-simple vector and subcloned into expression vector pET28a. The prokaryotic expression plasmid pET28a-sao was transformed into E. coli BL21 after the identification with double digestion of BamH I and Sal I and DNA sequencing. With the induced expression by isopropyl fl-D-l-thiogalacto- pyrannoside（IPTG） ,the recombinant protein Sao was analyzed by sodium dodecyl sulfate polyacrylamide gel electro- phresis （SDS-PAGE）. The Sao protein was purified with His-Binding-resin. The titer and specificity of the polyclonal an- tibodies were determined by enzyme-linked immunosorbent assay（ELISA） and Western blot. Bactericidal assay was used to identify the bactericidal activity of rabbit polyclonal antibodies against Sao. Results The expression vector pET28a- sao was successfully constructed. The fused gene was highly-expressed in E. coli BL21. ELISA detection results indicated that the titer of polyclonal antibodies to Sao was about 1 ： 102400. Western blot results showed that the anti-Sao polyclonal antibodies could bind to recombinant Sao protein specifically. The polyclonal antibodies against the recombinant Sao pro- tein could promote bactericidal ability of human whole-blood. Conclusion The anti-Sao polyclonal antibodies are of good specificity and high titer and could promote bactericidal ability of human whole-blood.