title: [Expression and purification of Legionella pneumophila MIP protein and its application in serological diagnosis].
abstract: OBJECTIVE
  To express and purify macrophage infectivity potentiator (MIP) protein of Legionella pneumophila(Lp), and explore its value in the serological diagnosis of Lp.
  
  
  METHODS
  The recombinant plasmid pET-mip was transformed into E.coli BL21 competent cells. The expression of MIP protein was induced, and then analyzed by SDS-PAGE electrophoresis, purified by affinity chromatography. We screened out 40 positive blood serum and 30 negative blood serum using the DRG (Germany, IgG/IgM/IgA) Lp kit. The blood serum samples were detected for IgG, IgM, IgA antibody levels by indirect ELISA that we had established with the purified MIP protein as the coating antigen, as well as by R&D (USA, IgG/IgM/IgA) Lp kit. The two methods were compared in the sensitivity, specificity and consistency of the test results.
  
  
  RESULTS
  The recombinant MIP protein was successfully expressed and purified with Mr; being 40 000 in E.coli BL21. In comparison of the indirect ELISA we developed with the R&D Lp kit for detecting Lp antibody IgG, IgM and IgA in blood serum, the specificity of IgG was 88.5% and the sensitivity was 95.5%, the Kappa value was 0.846 (P<0.05), the area under the ROC curve was 0.927; the specificity of IgM was 89.3% and the sensitivity was 97.6%, the Kappa value was 0.88 (P<0.05), the area under the ROC curve was 0.947; the specificity of IgA was 90% and the sensitivity was 95.2%, the Kappa value was 0.856 (P<0.05), the area under the ROC curve was 0.931.
  
  
  CONCLUSIONS
  MIP proteins of L.pneumophila was expressed and purified successfully, and MIP protein can be used as a coating antigen in serological diagnosis of L.pneumophila.
authors.affiliation: Department of Pathogenic Biology and Immunology, School of Basic Medicine, Ningxia Medical University, Ministry of Education, Yinchuan 750004, China.
authors.forename: Li
authors.initials: L
authors.lastname: Liu
authors.forename: Xiuqin
authors.initials: X
authors.lastname: Cao
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Yang
mesh_terms: D000907:Antibodies, Bacterial / Q000097:blood / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D006801:Humans; D016952:Legionella pneumophila / Q000235:genetics* / Q000276:immunology; D007877:Legionnaires' Disease / Q000175:diagnosis*; D019696:Peptidylprolyl Isomerase / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D012372:ROC Curve; D011933:Reagent Kits, Diagnostic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification
chemical_list: D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D011933:Reagent Kits, Diagnostic; D011993:Recombinant Fusion Proteins; C512922:Mip protein, Legionella pneumophila; D019696:Peptidylprolyl Isomerase