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纸质出版:2014
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闵迅,黄健,黄美容,钟文,董杰,陈特,张雪梅.肺炎链球菌蛋白PotD的表达纯化及免疫活性分析[J].陆军军医大学学报(原第三军医大学学报),2014,36(16):1660-1663.
Min Xun,Huang Jia,Huang Meirong,Zhong Wen,Dong Jie,Chen Te,Zhang Xuemei.Expression, purification and immunological activity analysis of PotD protein from Streptococcus pneumoniae[J].J Amry Med Univ (J Third Mil Med Univ),2014,36(16):1660-1663.
闵迅,黄健,黄美容,钟文,董杰,陈特,张雪梅.肺炎链球菌蛋白PotD的表达纯化及免疫活性分析[J].陆军军医大学学报(原第三军医大学学报),2014,36(16):1660-1663. DOI:
Min Xun,Huang Jia,Huang Meirong,Zhong Wen,Dong Jie,Chen Te,Zhang Xuemei.Expression, purification and immunological activity analysis of PotD protein from Streptococcus pneumoniae[J].J Amry Med Univ (J Third Mil Med Univ),2014,36(16):1660-1663. DOI:
目的 研究PotD蛋白免疫保护活性,探讨其作为疫苗候选蛋白的可能性。 方法 构建、表达、纯化PotD蛋白,通过免疫BALB/c小鼠以制备多克隆抗体,评估其免疫原性,并进一步分析其对儿童的免疫保护效应;采用PCR技术扩增8株不同血清型
S.pn
中PotD基因后进行测序鉴定分析基因序列的保守性;再通过western blot技术分析8株菌中PotD蛋白的表达水平,以评估PotD蛋白保守性;采用粘附抑制实验分析PotD蛋白的免疫保护活性。 结果 成功获得纯度大于95%的PotD目的蛋白,将其免疫小鼠后获得高效价抗体;在10岁以下儿童
S.pn
感染者及健康带菌者血清中均有抗PotD存在;对8株不同血清型
S.pn
PotD基因进行PCR扩增和测序发现8株细菌中该基因具有高度保守性;Western blot技术分析证实,8株不同血清型
S.pn
均表达该蛋白;粘附抑制实验证实,重组PotD蛋白及其抗体均能够显著抑制
S.pn
对A549细胞的粘附作用。 结论 PotD蛋白制备方便,在我国流行
S.pn
血清型中有良好的保守性和免疫原性,能够显著抑制
S.pn
对宿主细胞的粘附,可能成为良好的蛋白疫苗靶点。
Objective To study the immunoprotective activity of PotD proteins from
Streptococcus pneumoniae
(
S.pn
) and its application possibility as a vaccine candidate. Methods After expression and purification of PotD protein
BALB/c mice were immunized with PotD protein to produce polyclonal antibody. The immunogenicity of PotD protein was assessed
and then further its effects on children were analyzed. The conservation of PotD gene in 8 different
S.pn
strains was tested by P
CR and sequencing. Using Western blot
the expression level and the conservation of PotD protein in 8 different
S.pn
strains was tested. Adherence inhibition experiments were used to analyze the immunogenicity of PotD protein. Results Recombinant PotD protein with high purity and high-titer anti-PotD antibody were successfully obtained. Anti-PotD antibody was also found in the sera from children under the age of 10 who were infected with
S.pn
. The PotD gene was highly conservative in the 8 different
S.pn
strains. All of the 8
S.pn
strains expressed the PotD protein. Adherence inhibition experiments showed that the recombinant PotD and anti-PotD antiserum had a significant inhibition effect on
S.pn
adhesion to A549 cells. Conclusion PotD protein with high conservation and immunological activity can be prepared conveniently. It can significantly inhibit
S.pn
adhesion to host cells and may be a good protein vaccine candidate.
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