摘要
目的表达LTB-NspA重组融合蛋白,鼻饲免疫雌性Balb/c小鼠,分析重组融合蛋白的免疫活性,为研制抗淋球菌蛋白疫苗提供实验依据。方法转化并鉴定pET30a/ltB-nspA原核重组质粒后,在E.coliBL21中表达重组融合蛋白。鼻饲途径免疫雌性Balb/c小鼠,检测NspA特异性体液免疫和细胞免疫水平。结果 LTB-NspA融合蛋白组生殖道黏膜产生的NspA特异性sIgA水平和血清IgG水平随免疫时间呈上升趋势,第6周sIgAA450nm达0.689,明显高于NspA组和LTB、蛋白溶解液(solution buffer)对照组(P<0.01);第6周血清中产生的特异性IgGA450nm值达0.734,明显高于LTB和Solution Buffer对照组(P<0.01);LTB-NspA组脾淋巴细胞刺激指数(SI)为1.63,明显高于LTB和Solution Buffer对照组(P<0.01);LTB-NspA组小鼠脾淋巴细胞经特异性抗原NspA刺激后,培养上清中IFN-γ和IL-4含量明显升高,分别达142.23pg/ml和86.14pg/ml,明显高于LTB和Solution Buffer对照组(P<0.01)。但LTB-NspA与NspA两组间血清IgG水平、SI、IFN-γ和IL-4水平差异不显著(P>0.05)。结论 LTB-NspA重组蛋白诱导小鼠产生了较高水平的特异性体液免疫和细胞免疫;首次证实黏膜佐剂LTB可辅佐NspA诱导小鼠产生高水平的生殖道黏膜免疫。
The aim of the study is to express LTB-NspA fusion protein and identify immunocompetence of the protein,and thus provide excremental basis for preparing anti-gonorrhoeae protein vaccine.The prokaryotic recombinant plasmid pET-30a/ltB-nspA was extracted,and then identified by PCR and the double-enzymes digestion,finally transformed into E.coli BL21 for protein expression.We analyzed the immunocompetence（humoral immunologic response and the cellular immunologic response） of the recombinant fusion protein in female Balb/c mice through intranasally immunization.The levels of NspA-specific sIgA in murine genital tract and IgG in serum showed upward trend along with the weeks post innoculation in LTB-NspA group.A450nm of sIgA in LTB-NspA group was 0.689 at the week 6,which was significantly higher than the group NspA,LTB and solution buffer controls（P 0.01）.A450 nm of IgG（0.734） at the week 6,stimulation index（1.63） of the splenic lymphocyte,the levels of IFN-γ（142.23 pg/ml） and IL-4（86.14 pg/ml） induced by splenic lymphocyte in LTB-NspA group were significantly higher than those of the LTB and solution buffer controls（P 0.01）.However,the levels of IgG,SI,IFN-γ,and IL-4 between LTB-NspA group and NspA group had no significant differences（P 0.05）.All these results indicate that the recombinant fusion protein LTB-NspA could induce high level of humoral immunoresponse and slightly cellular immunologic response in female Balb/c mice through intranasally immunization.Furthermore,we found the mucosal adjuvant LTB could assist NspA to induce higher level of mucosal immune response in the genital tract mucosa of mice.
出处
《免疫学杂志》
CAS
CSCD
北大核心
2010年第11期982-986,共5页
Immunological Journal
基金
国家自然科学基金(30771931)
湖南省教育厅科研基金(08C745)
关键词
淋病奈瑟菌
奈瑟菌表面蛋白A
LTB
重组蛋白
免疫活性
Neisseria gonorrhoeae
Neisseria surface protein A
LTB
Recombinant protein
Immunocompetence
