title: Leptospirosis serodiagnosis by ELISA based on recombinant outer membrane protein.
abstract: The outer membrane protein LipL21, LipL32, LipL41 and Loa22 of Leptospira interrogans serovar Copenhageni were previously revealed by immunoproteomic analysis, using sera from acute phase infection in a guinea pig. The full-length DNA of each protein was then cloned from the same serovar and expressed in pRSET vector. The obtained molecular weight (MW) of recombinant proteins rLipL21, rLipL32 and rLoa22 were slightly higher than the MW predicted from nucleotide sequences of each inserted gene, while only the N-terminal half of rLipL41 was obtained. Mice antiserum raised against each purified recombinant protein could react with the whole cell lysate of leptospiral serovars, implying that leptospiral native proteins shared a common epitope with recombinant protein. Serodiagnosis using recombinant protein antigen based on indirect ELISA procedure was developed in this study. The optimization of the ELISA components lead to determination of optical density (OD) from a single serum-dilution of 1:1000 in the leptospirosis patients group and normal healthy control group. The cut off OD values for both IgG and IgM class were investigated, and based on this fixed dilution only the IgG class could be used for differential diagnosis of patients and normal individuals. Compared with the MAT assay, ELISA assay utilizing both rLipL32 and rLoa22 as antigen, gave high accuracy and could thus be useful as a confirmative serology test.
authors.affiliation: Department of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Piyanart
authors.initials: P
authors.lastname: Chalayon
authors.forename: Phanita
authors.initials: P
authors.lastname: Chanket
authors.forename: Toungporn
authors.initials: T
authors.lastname: Boonchawalit
authors.forename: Siriporn
authors.initials: S
authors.lastname: Chattanadee
authors.forename: Potjanee
authors.initials: P
authors.lastname: Srimanote
authors.forename: Thareerat
authors.initials: T
authors.lastname: Kalambaheti
mesh_terms: D000372:Agglutination Tests / Q000379:methods; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood*; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D006168:Guinea Pigs; D007074:Immunoglobulin G / Q000097:blood*; D007075:Immunoglobulin M / Q000097:blood*; D007921:Leptospira interrogans / Q000276:immunology* / Q000302:isolation & purification; D007922:Leptospirosis / Q000175:diagnosis*; D008074:Lipoproteins / Q000235:genetics / Q000276:immunology; D008297:Male; D051379:Mice; D010802:Phylogeny; D011994:Recombinant Proteins / Q000235:genetics; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D007074:Immunoglobulin G; D007075:Immunoglobulin M; C545585:LipL21 protein, Leptospira interrogans; C406589:LipL32 protein, Leptospira; C538745:LipL41 protein, Leptospira interrogans; D008074:Lipoproteins; D011994:Recombinant Proteins