| 洪文艳1，2 王浩然1 王津1 郭兆彪1 周蕾1 杨瑞馥1.鼠疫菌PsaA 抗原的表达及抗体制备和应用[J].,2011,11(12):2201-2203 |
| 鼠疫菌PsaA 抗原的表达及抗体制备和应用 |
| PsaA Antigen Expression of Y. pestis and Its Antibody Developmentand Application |
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| DOI： |
| 中文关键词: 鼠疫菌 PsaA 抗原 蛋白表达 抗体制备 抗体检测 |
| 英文关键词: Y. pestis PsaA antigen Protein expression Antibody development Antibody detection |
| 基金项目:国家973 项目（2007CB310505）；国家科技支撑计划项目（2009BAK43B31）；北京市自然科学基金（5082017） |
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| 中文摘要: |
| 目的: 制备鼠疫菌PsaA 抗原及抗体，建立针对PsaA 抗体的快速检测方法，并检测鼠疫感染猴血清标本中的PsaA 抗体的阳
性率。方法：利用PCR 方法扩增出PsaA 蛋白基因片段，在大肠杆菌原核表达系统中表达出重组PsaA 抗原，以镍柱亲和层析纯化
包涵体形式的表达蛋白，以尿素梯度透析复性成可溶蛋白。再以表达蛋白为免疫原，常规免疫家兔，收集兔血清制备多抗，并以正
辛酸- 硫酸铵法提纯获得PsaA 抗体IgG。利用得到的PsaA 抗原和抗体为材料，建立两种检测PsaA 抗体的快速检测方法，即间接
ELISA 法和上转换发光(Up-converting Phospher Technology, UPT)免疫层析试纸条法。最后利用这两种方法检测18 份猴血清标本
中的PsaA 抗体。结果：鼠疫菌感染猴血清标本中PsaA 抗体的阳性率为62%(8/13)。结论：成功建立了针对鼠疫菌PsaA 抗体的快
速检测方法，并检测到13 份鼠疫感染猴血清标本中的PsaA 抗体的阳性率为62%。 |
| 英文摘要: |
| Objective: To develop the PsaA antigen of Y.Pestis and its antibody, and to develop the fast detection assays, and to
detect the positive ratio of PsaA antibody in 18 plague monkey serum samples. Methods: The PsaA gene was amplificated by PCR
method. The production was cloned and expressed using an E. coli expression system, purified with the Ni-NTA affinity chromatography,
and renatured the inclusion body expressed proteins using a urea gradient dialysis method. The corresponding polyclonal antibody was
prepared by immunizing rabbits using the conventional method. Bloods were obtained from rabbits for collection of sera and purification
of specific immunoglobulin G (IgG) with a caprylic acid-saturated ammonium sulfate precipitation method. Then both indirect ELISA
method and Up-converting Phospher Technology (UPT) lateral flow assay were developed to detect PsaA antibodies of samples. Lastly,
18 monkey serum samples were tested. Results: The data showed that the positive ratio of the PsaA antibody in plague monkey serum
samples was 62%(8/13). Conclusion: The fast detection assays were successfully developed for the PsaA antibody test. And the positive
ratio of the antibody in plague monkey serum samples was 62%. |
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