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宋内氏痢疾杆菌 (S .Sonnei)的一个重要保护性抗原是菌体抗原 (O抗原 )即细菌细胞壁中的脂多糖 (LPS)成分 ,已证实血清中足够水平的抗LPSIgG抗体即能对该菌提供免疫保护作用。本文选用Westphal热酚法提纯宋内氏痢疾菌LPS ,经酸水解法进一步脱毒处理后得到无毒但却具弱免疫原性的O 特异性多糖 (O SP) ,然后采用化学方法即通过连接剂己二酸二肼 (ADH)将其共价结合到破伤风类毒素 (TT)上 ,共制备得到了三批宋内氏痢疾杆菌O SP TT结合疫苗。经生化和免疫学方法检测证实我们提纯的LPS、OSP及其合成的O SP AH衍生物、O SP TT结合物均具有宋内氏痢疾杆菌O抗原特异性 ,同时其核酸和杂蛋白质含量低 (≤ 2 % ) ,表明纯度较高。同时经小鼠免疫原性试验证实三批结合物免疫小鼠后产生的抗LPSIgG抗体滴度均比单一O SP免疫后产生的要高出 10倍以上 ,且结合物再次注射后存在加强应答。补体介导的体外杀菌力试验表明结合物免疫小鼠后诱导产生的血清抗LPSIgG抗体对宋内氏痢疾杆菌具特异性杀菌活性。本文结果表明宋内氏痢疾杆菌O SP TT结合疫苗可望作为一种有效的侯选痢疾疫苗做进一步的大规模人体观察。
An important protective antigen of S. sonnei is the bacterial antigen (O antigen), a component of lipopolysaccharide (LPS) in the cell wall of bacteria, which has been shown to be sufficient against serum LPS IgG antibodies Bacteria provide immune protection. In this paper, Westphal thermal phenolic method was used to purify LPS from S.aeruginosa, and the nontoxic but weakly immunogenic O-specific polysaccharide (O SP) was obtained after further detoxification by acid hydrolysis and then chemically linked to A total of three batches of Shigella dysenteriae O SP TT conjugate vaccine were co-prepared by covalent attachment of ADH to tetanus toxoid (TT). Biochemical and immunological methods confirmed that our purified LPS, OSP, and their synthesized O SP AH derivatives, O SP TT conjugates, all have the specificity of Shigella sonnei O antigen with low levels of nucleic acids and heteroplasmas ≤ 2%), indicating higher purity. At the same time, the mice immunogenicity test confirmed that three batches of conjugate immunized mice produced anti-LPS IgG antibody titers than the single O SP immunization produced more than 10 times higher, and the combination of re-injection there is a strong response. Complement-mediated in vitro bactericidal tests showed that the anti-LPS IgG antibody produced by the conjugate immunized mice showed specific bactericidal activity against Shigella flexneri. Our results show that Shigella sonnei O SP TT conjugate vaccine is expected to be used as an effective candidate dysentery vaccine for further large-scale human observation.