Production of recombinant Bartonella henselae 17-kDa protein for antibody-capture enzyme-linked immunosorbent assay

Diagn Microbiol Infect Dis. 2006 May;55(1):1-7. doi: 10.1016/j.diagmicrobio.2005.10.020. Epub 2006 Feb 20.

Abstract

The Bartonella henselae 17-kDa protein was expressed in a prokaryotic expression system as a histidine-tagged fusion protein and was purified. The target gene was cloned into a recombinant expression construct, pTri-17kd. The expressed protein was purified to near homogeneity by a nickel-agarose column chromatography. Protein recovery was estimated to be 2.9 mg from 100 mL of bacterial culture. The purified 17-kDa protein was recognized by serum from patients infected with B. henselae and Bartonella quintana, suggesting antigenic integrity. The sensitivity and specificity of the IgG enzyme-linked immunosorbent assay (ELISA) relative to immunofluorescent antibody assay testing were 71.1% and 93.0%, respectively. According to the receiver operating characteristic curve analysis, the area under the curve was 0.823. These results indicate that the expressed 17-kDa protein is a suitable source of antigen for development of an antibody-capture ELISA for the detection of antibodies to B. henselae.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antibodies, Bacterial / blood*
  • Antigens, Bacterial / immunology*
  • Area Under Curve
  • Bacterial Proteins / genetics
  • Bacterial Proteins / immunology*
  • Bartonella henselae / genetics
  • Bartonella henselae / immunology*
  • Bartonella quintana / genetics
  • Bartonella quintana / immunology
  • Blotting, Western
  • Cats
  • Chromatography, Agarose
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme-Linked Immunosorbent Assay / methods*
  • Escherichia coli / genetics
  • Fluorescent Antibody Technique, Indirect
  • Gene Amplification
  • Gene Expression Regulation / immunology*
  • Genetic Vectors
  • Humans
  • Immunoglobulin G / analysis
  • Molecular Sequence Data
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology*

Substances

  • Antibodies, Bacterial
  • Antigens, Bacterial
  • Bacterial Proteins
  • Immunoglobulin G
  • Recombinant Proteins