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Preparation of Two Different Lengths of FopA Antigen and Antibody Used in Francisella tularemia Detecting |
JING Ying-ying1,2, YANG Yu1, WANG Jing1, YANG Yong-li1, HU Kong-xin1, WANG Zhen-dong2 |
1. Chinese Academy of Inspection and Quarantine, Beijing 100123, China;
2. College of Bioscience and Biotechnology, Shenyang Agricultural University, Shenyang 110161, China |
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Abstract Objective:To establish an BL21 expression system of Francisella tularemia-specific antigen (FopA-L and FopA-S), then obtain their highly active recombinant protein and the polyclonal antibodies which can be applied for Francisella monitoring, diagnosis and treatment.Methods:Constructing a plasmid expression vector pET100 FopA-L and FopA-S, then transforming FopA plasmid into E. coli BL21 cells, induced by IPTG, purifing protein by chelate nickel ion-NTA (Ni-NTA) affinity chromatography, making polyclonal antibody by recombinant FopA-L and FopA-S proteins immune rabbits and antibody were determined by Western blot, ELISA and GICA. Results:Constructed expression vectors pET100 of FopA-L and FopA-S, the BL21 cell lines obtained can highly express target proteins, antibody specifically combined with them which can be used in Francisella detecting were prepared successfully, with a titer of more than 1 ∶100000 and highly specific. Conclusion:Preparation the FopA-L and FopA-S antigen and antibody laid the foundation for the establishment of rapid detection method of Francisella.
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Received: 06 August 2010
Published: 25 December 2010
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[1] |
McLendon M K, Apicella M A, Allen L A. Francisella tularensis: taxonomy, genetics, and Immunopathogenesis of a potential agent of biowarfare. Annu Rev Microbiol, 2006, 60: 167-185.
|
|
|
[2] |
Hopla C E. The ecology of tularemia.Adv Vet Sci Comp Med, 1974, 18: 25-53.
|
|
|
[3] |
宫英, 逄增昌, 强新,等. 土拉弗氏菌病患者血清抗体20年变化趋势分析. 中国公共卫生, 2006, 22(10): 1201-1202. Gong Y, Pang Z C, Qiang X, et al. Chinese Journal of Public Health, 2006, 22(10): 1201-1202.
|
|
|
[4] |
Porsch-Ozcurumez M, Kischel N, Priebe H, et al. Comparison of enzyme-linked immunosorbent assay, Western blotting, microagglutination, indirect immunofluorescence assay, and flow cytometry for serological diagnosis of tularaemia. Clinical and Diagnostic, 2004, 11(6): 1001-1015.
|
|
|
[5] |
Aronova N V, Pavlovich N V. The use if lipopolysaccharide in the dot solid phase enzyme immunoassay. Mikrobiol Epidemiol Immunobiol, 2000, 5: 75-78.
|
|
|
[6] |
Berdal B P, Mehl R, Haaheim H, et al. Field detection of Francisella tularensis.Scand J Infect Dis, 2000, 32(3): 287-291.
|
|
|
[7] |
Magnarelli L, Levy S, Koski R. Detetion of antibodies to Francisella tularensis in cats.Rvsc, 2007, 82(1): 22-26.
|
|
|
[8] |
Tarnvik A, Berglund L. Tularemia. Eur Respir J, 2003, 21(2): 361-373.
|
|
|
[9] |
靖学芳, 安云庆. LPS和抗LPS治疗的研究及应用进展. 微生物学免疫学进展, 2004, 32(2): 53-57. Jing X F, An Y Q. Progress in Microbiology and Immunology, 2004, 32(2): 53-57.
|
|
|
[10] |
Huntley J F, Conley P G, Hagman K E, et al. Characterization of Francisella tularensis outer membrane proteins. J Bacteriol, 2007, 189(2): 561-574.
|
|
|
[11] |
Fulop M, Manchee R, Titball R. Role of lipopolysaccharide and a major outer membrane protein from Francisella tularensis in the induction of immunity against tularemia. Vaccine, 1995, 13(13):1220-1225.
|
|
|
[12] |
Saier M H, Werner P K, Muller M. Insertion of proteins into bacterial membranes: mechanis, characteristics, and comparisons with the eucaryotic process. Microbiol Rev, 1989, 53(3): 333-366.
|
|
|
[13] |
Gunnar von Heijne. Life and death of a signal peptide. Nature, 1998, 396 (12):111-113.
|
|
|
[14] |
何斌, 端青. 重组土拉弗朗西斯菌外膜蛋白 FopA 的融合表达及抗原性分析. 生物技术通讯, 2010, 21(1): 32-34. He B, Duan Q. Letters in Biotechnology, 2010, 21(1): 32-34.
|
|
|
[15] |
王增, 马会勤, 张文,等. 包涵体蛋白的分离和色谱法体外复性纯化研究进展. 中国生物工程杂志, 2009, 29(7): 102-107. Wang Z, Ma H Q, Zhang W, et al.China Biotechnology, 2009, 29(7): 102-107.
|
|
|
[16] |
张婷婷, 叶波平. 包涵体蛋白质的复性研究进展. 药物生物技术, 2007, 14 (4): 306-309. Zhang T T, Ye B P. Pharmaceutical Biotechnology, 2007, 14 (4): 306-309.
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