title: Validation of an improved competitive enzyme-linked immunosorbent assay to detect Equine arteritis virus antibody.
abstract: The objective of the present study was to validate a previously described competitive enzyme-linked immunosorbent assay (cELISA) to detect antibody to Equine arteritis virus (EAV) based on GP5-specific nonneutralizing monoclonal antibody (mAb) 17B7(9) using the World Organization for Animal Health (OIE)-recommended protocol, which includes the following 5 in-house analyses. 1) The assay was calibrated with the OIE-designated reference serum panel for EAV; 2) repeatability was evaluated within and between assay runs; 3) analytical specificity was evaluated using sera specific to related viruses; 4) analytical sensitivity was evaluated with sera from horses vaccinated with an EAV modified live virus (MLV) vaccine; and 5) the duration of cELISA antibody detection following EAV vaccination was determined. The positive cELISA cutoff of ≥35% inhibition (%I) was confirmed by receiver operating characteristic plot analysis. Analytical sensitivity of the cELISA was comparable to the serum neutralization (SN) assay in that it detected EAV-specific antibody as early as 8 days postvaccination. The duration of EAV-specific antibody detected by cELISA was over 5 years after the last vaccination. This cELISA could detect EAV-specific antibody in serum samples collected from horses infected with various EAV strains. In the field trial performed by American Association of Veterinary Laboratory Diagnosticians-accredited state laboratories and OIE laboratory, the diagnostic specificity of the cELISA was 99.5% and the diagnostic sensitivity was 98.2%. The data using various serum panels also had consistently significant positive correlation between SN titers and cELISA %I results. The results further confirm that the EAV antibody cELISA is a reliable, simple alternative to the SN assay for detecting EAV-specific antibodies in equine sera.
authors.affiliation: 1Chungwon Chung, VMRD (Veterinary Medical Research and Development) Inc., Pullman, WA 99163. chungwon@vmrd.com.
authors.forename: Chungwon
authors.initials: C
authors.lastname: Chung
authors.forename: Carey
authors.initials: C
authors.lastname: Wilson
authors.forename: Peter
authors.initials: P
authors.lastname: Timoney
authors.forename: Udeni
authors.initials: U
authors.lastname: Balasuriya
authors.forename: Ethan
authors.initials: E
authors.lastname: Adams
authors.forename: D Scott
authors.initials: DS
authors.lastname: Adams
authors.forename: James F
authors.initials: JF
authors.lastname: Evermann
authors.forename: Alfonso
authors.initials: A
authors.lastname: Clavijo
authors.forename: Kathleen
authors.initials: K
authors.lastname: Shuck
authors.forename: Sandy
authors.initials: S
authors.lastname: Rodgers
authors.forename: Stephen Sauchi
authors.initials: SS
authors.lastname: Lee
authors.forename: Travis C
authors.initials: TC
authors.lastname: McGuire
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral / Q000097:blood*; D018174:Arterivirus Infections / Q000097:blood / Q000662:veterinary* / Q000821:virology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D007240:Equartevirus / Q000302:isolation & purification*; D006734:Horse Diseases / Q000097:blood / Q000821:virology*; D006736:Horses; D009500:Neutralization Tests / Q000662:veterinary; D012372:ROC Curve; D015203:Reproducibility of Results; D018709:Statistics, Nonparametric
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral
keywords: Antibody; Equine arteritis virus; assay validation; competitive enzyme-linked immunosorbent assay; serum neutralization