Development and evaluation of a MAb based competitive-ELISA using helicase domain of NS3 protein for sero-diagnosis of bovine viral diarrhea in cattle and buffaloes.
The aim of this study was to develop a competitive inhibition ELISA (CI-ELISA) for detection of antibodies to bovine viral diarrhea virus (BVDV) using the helicase domain of NS3 (non-structural) protein and monoclonal antibody (MAb) against it and to estimate its sensitivity and specificity using two commercial ELISA kits as independent references. The 45-kDa helicase domain of NS3 protein of BVDV was expressed in Escherichia coli and 18MAbs were developed against it. MAb-11G8 was selected for use in CI-ELISA on the basis of maximum inhibition (90%) obtained with BVDV type 1 infected calf serum. Based on the distribution of percent inhibition of known negative sera (n=166), a cut-off value was set at 40% inhibition. In testing 914 field serum samples of cattle (810) and buffaloes (104), the CI-ELISA showed a relative specificity of 95.75% and 97.38% and sensitivity of 96% and 94.43% with Ingenesa kit and Institut Pourquier kit, respectively. This study proved that the use of helicase domain of NS3 (45-kDa) is equally good as the whole NS3 protein (80-kDa) used in commercial kits for detection of BVDV antibodies in cattle and buffaloes.
Bhatia
S
S
High Security Animal Disease Laboratory, Indian Veterinary Research Institute, Anand Nagar, Bhopal 462 021, Madhya Pradesh, India. bhatia_san@yahoo.co.in
Sood
Richa
R
Mishra
N
N
Pattnaik
B
B
Pradhan
H K
HK
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000276:immunology; D001912:Bovine Virus Diarrhea-Mucosal Disease / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D002020:Buffaloes; D002417:Cattle; D001908:Diarrhea Viruses, Bovine Viral / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000592:standards / Q000662:veterinary*; D010447:Peptide Hydrolases / Q000276:immunology*; D020365:RNA Helicases / Q000276:immunology*; D012680:Sensitivity and Specificity; D017361:Viral Nonstructural Proteins / Q000276:immunology*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D010447:Peptide Hydrolases; D020365:RNA Helicases; D017361:Viral Nonstructural Proteins; C086730:p80 protein, bovine viral diarrhea virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases