Evaluation of a commercial ELISA test kit on classical swine fever antibody detection using oral fluid samples

Abstract

Classical swine fever virus (CSFV) contributes to economic loss of swine production in endemic countries.
Serum neutralization and enzyme-linked immunosorbent assay (ELISA) are serological tests commonly used for
monitoring CSFV antibody status with serum samples. This experiment evaluated the detection of CSFV antibody in
oral fluid samples in commercial ELISA test kit by in vitro study of negative oral fluid mixed with serum of known
CSFV serum neutralizing (SN) titer and in vivo oral fluid samples obtained from experimental animals. Correlations of
SN titer and S/P ratio of ELISA were observed with the in vitro oral fluid samples, indicating the stability of the antibody
in the oral fluid and the potential of oral fluid as an alternative specimen for CSFV diagnosis. Diagnostic sensitivity of
the oral fluid detection using in vitro samples was as high as 95.83% when the ELISA procedures were modified (i.e. 12
h incubation at 4°C and addition of 100 µl antibody conjugate). The in vivo experiment used oral fluid and blood samples
obtained from 20 piglets (20 days old) which were divided into 3 experimental groups: challenged with ALD strain,
low virulence CSFV (A) (n=8); vaccinated (B) (n=8); and negative control (C) (n=4). The animals were vaccinated with
CSFV LOM strain modified live vaccine at 0 day post inoculation (dpi) and re-challenged (A&B) with high virulence
CSFV on 14 dpi and later euthanized at 30 dpi. Results from the in vivo oral fluid samples demonstrated low detectable
antibody titers (SN titer 0-4) using neutralizing peroxidase-linked assay (NPLA) and all samples were negative to CSFV
indirect ELISA performed at the optimal condition obtained from the in vitro protocol.