Dot-enzyme immunoassay for visual detection of antibodies to pseudorabies virus in swine serum

Abstract

A modified solid-phase enzyme immunoassay (EIA) is described for the visual detection of anti-pseudorabies virus (anti-PRV) antibody in porcine serum. Dots of PRV antigens were adsorbed to nitrocellulose paper (hence the name dot-EIA), and the remaining nonspecifically reactive sites were blocked with bovine serum albumin or skim milk powder. After immersion in test serum, bound antibodies were reacted with a peroxidase-conjugated anti-porcine immunoglobulin G (H & L). Positive reactions were easily visualized as brown dots after enzyme degradation of a substrate containing hydrogen peroxide and diaminobenzidine. The dot-EIA was comparable to the serum neutralization test and the standard microtiter EIA in its ability to detect antibody in the sera of pigs 9 days after experimental infection and 12 days after contact with infected pigs. The sensitivity and specificity of the dot-EIA relative to the serum neutralization test and the standard EIA were determined from the testing of 856 field sera from the United Kingdom, the United States, and Canada. In all comparisons, both the relative sensitivity and specificity of the dot-EIA were in the order of 98 to 99%. The dot EIA appears to have potential application as a rapid and economical field test in the diagnosis of PRV infection.