Standardization of enzyme-linked immunosorbent assays (ELISAs) for quantitative estimation of antibodies specific for infectious bovine rhinotracheitis virus, respiratory syncytial virus, parainfluenza-3 virus, and bovine viral diarrhea virus.
Commercial enzyme-linked immunosorbent assays (ELISAs) for detection of serum antibodies to bovine viral diarrhea virus (BVDV), parainfluenza-3 virus (PI3V), respiratory syncytial virus (RSV), and infectious bovine rhinotracheitis virus (IBRV) were standardized to give a quantitative result when testing was performed at a single optimum dilution. For each test, serum samples were titrated and their end point titers calculated by an algebraic method directly from a plot of each titration series and also from a regression line fitted to this plot. The corrected optical density (COD) of each sample when tested at dilutions of 1/25, 1/50, and 1/100 was expressed as a percentage of the COD of a positive reference serum included on each plate, this value was the sample/positive (S/P) ratio. For each test, the linear relationship between the S/P ratio obtained at a dilution of 1/25, 1/50, and 1/100 and the end point titer calculated by each method was determined. In each case, the best linear relationship existed when samples were tested at a dilution of 1/100 (r = 0.973 for BVDV, 0.962 for PI3V, 0.961 for RSV, 0.947 for IBRV). From the equation of these lines, an increase in the S/P ratio between acute and convalescent serum samples of 31%, 23%, 21%, and 35% would correspond to a 4-fold rise in ELISA titer to BVDV, PI3V, RSV, and IBRV, respectively. ELISA titers calculated from S/P ratios at 1/100 were significantly related to virus neutralization titers to BVDV, RSV, and IBRV and to hemagglutination inhibition titers to PI3V (P < < 0.001 in all cases). Samples with low S/P ratios had the greatest intraassay and interassay variation. Intraassay reproducibility ranged from 3.5% to 22.3% (coefficient of variation), with a median value of 9.5%. Interassay reproducibility was lower, ranging from 6.0% to 50.6%, with a median of 17.4%.
Graham
D A
DA
Veterinary Sciences Division, Department of Agriculture for Northern Ireland, Belfast.
Mawhinney
K A
KA
McShane
J
J
Connor
T J
TJ
Adair
B M
BM
Merza
M
M
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D001912:Bovine Virus Diarrhea-Mucosal Disease / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D002417:Cattle; D002418:Cattle Diseases; D001908:Diarrhea Viruses, Bovine Viral / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000592:standards / Q000662:veterinary; D007242:Herpesvirus 1, Bovine / Q000276:immunology; D007241:Infectious Bovine Rhinotracheitis / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D011786:Quality Control; D012044:Regression Analysis; D015203:Reproducibility of Results; D018357:Respiratory Syncytial Virus Infections / Q000175:diagnosis / Q000276:immunology / Q000662:veterinary*; D012136:Respiratory Syncytial Viruses / Q000276:immunology; D018108:Respirovirus / Q000276:immunology; D010253:Respirovirus Infections / Q000175:diagnosis / Q000276:immunology / Q000662:veterinary*; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral