title: A novel 12.6-kDa protein of Leishmania donovani for the diagnosis of Indian visceral leishmaniasis.
abstract: BACKGROUND
  For the diagnosis of visceral leishmaniasis (VL), rK39 antigen-based rapid test is widely used. Unfortunately, up to 32% healthy individuals from endemic region test positive with this antigen. There is an urgent need to search for a more specific antigen with sensitivity similar to rK39.
  
  
  METHODS
  We identified a Leishmania donovani-specific 12.6-kDa (BHUP3) soluble promastigote antigen through sensitive western blot technique. The identified protein was partially purified from sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the antigenic response of eluted protein was determined by western blot with different groups of individual sera. The diagnostic potential was further validated by enzyme-linked immunosorbent assay using serum of 100 VL patients, 93 nonendemic healthy control individuals, 110 endemic healthy control individuals, and 110 disease control individuals. Further, it was characterized by two-dimensional gel electrophoresis followed by matrix-assisted laser desorption/ionization-time-of-flight analysis.
  
  
  RESULTS
  On blotting, antibody against this protein was recognized by all (9/9) VL patient's sera, but it was absent in every control group (nonendemic healthy control and endemic healthy control). Sensitivity of the enzyme-linked immunosorbent assay was 88% (89/101), whereas the specificity for endemic healthy, nonendemic healthy, and different disease groups were 96% (106/110), 100% (93/93), and 97% (107/110), respectively. The two-dimensional gel electrophoresis showed a single spot, and matrix-assisted laser desorption/ionization-time-of-flight analysis revealed that it is a 113-amino-acid-long putative uncharacterized protein of 12.6-kDa anamorsin homolog matched completely with Leishmania major (GenBank accession number: Q4QIS1).
  
  
  CONCLUSIONS
  Despite marginally lower sensitivity of BHUP3, excellent specificity warrants its further development as a tool for diagnosis of VL.
authors.affiliation: Department of Medicine, Institute of Medical Sciences, Banaras Hindu University, Varanasi, Uttar Pradesh, India.
authors.forename: Dinesh
authors.initials: D
authors.lastname: Kumar
authors.forename: Subodh
authors.initials: S
authors.lastname: Kumar
authors.forename: Jaya
authors.initials: J
authors.lastname: Chakravarty
authors.forename: Shyam
authors.initials: S
authors.lastname: Sundar
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000913:Antibodies, Protozoan / Q000097:blood*; D000953:Antigens, Protozoan / Q000276:immunology / Q000302:isolation & purification; D015153:Blotting, Western; D019411:Clinical Laboratory Techniques / Q000379:methods*; D015180:Electrophoresis, Gel, Two-Dimensional; D019353:Endemic Diseases; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D006801:Humans; D007194:India; D007893:Leishmania donovani / Q000276:immunology*; D007898:Leishmaniasis, Visceral / Q000175:diagnosis* / Q000469:parasitology; D008297:Male; D008969:Molecular Sequence Data; D010691:Phlebotomus / Q000469:parasitology; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan