title: Development and evaluation of a novel antigen capture assay for the detection of classical swine fever virus antigens.
abstract: An antigen-capture enzyme immunoassay (EIA) was developed to detect classical swine fever virus (CSFV) antigen directly from 10% w/v tissue suspension. The assay, based on the sandwich principle, uses a biotinylated monoclonal antibody bound to streptavidin-coated microplates as the capture system and a swine anti-CSFV antibody and rabbit anti-swine HRPO-conjugate as the detector system. The antigen-capture EIA was compared with conventional virus isolation and polymerase chain reaction (PCR) for detection of CSFV in tissues. The ability of the antigen-capture EIA to discriminate classical swine fever (CSF) from bovine viral diarrhea and African swine fever viruses was also tested. The assay was shown to detect 21 different strains of CSFV and was unreactive with tissues from uninfected animals. Signal to noise (S/N) ratios were calculated from the EIA absorbance values. Readings from samples positive by virus isolation (n = 47) averaged a S/N ratio of 5.34. In contrast, samples negative by virus isolation (n = 96) demonstrated a mean S/N ratio of 0.16. At S/N cut-off value of 1.0, all samples that yield virus isolation and PCR negative result were negative in the antigen-capture EIA. Compared with virus propagation in tissue culture using PK15 cells (followed by indirect peroxidase assay detection) and PCR, the EIA had a specificity of 98.7% and a sensitivity of 91.4%. The EIA is simple, can be performed in 4 h and lends itself to automation for screening of tissues sample from pigs suspected of CSFV infection.
authors.affiliation: National Center for Foreign Animal Disease, Winnipeg, Manitoba, Canada. aclavijo@em.agr.ca
authors.forename: A
authors.initials: A
authors.lastname: Clavijo
authors.forename: E M
authors.initials: EM
authors.lastname: Zhou
authors.forename: S
authors.initials: S
authors.lastname: Vydelingum
authors.forename: R
authors.initials: R
authors.lastname: Heckert
mesh_terms: D000357:African Swine Fever / Q000175:diagnosis; D000358:African Swine Fever Virus / Q000302:isolation & purification; D000818:Animals; D000956:Antigens, Viral / Q000032:analysis*; D001912:Bovine Virus Diarrhea-Mucosal Disease / Q000175:diagnosis; D002417:Cattle; D002460:Cell Line; D006691:Classical Swine Fever / Q000175:diagnosis*; D006692:Classical Swine Fever Virus / Q000302:isolation & purification*; D003937:Diagnosis, Differential; D007124:Immunoenzyme Techniques; D010576:Pestivirus / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods; D011817:Rabbits; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013552:Swine
chemical_list: D000956:Antigens, Viral