Dot enzyme immunoassay for visual detection of peste-des-petits-ruminants virus antigen from infected caprine tissues.

摘要

An enzyme-linked immunosorbent microassay using nitrocellulose paper as the solid-phase support was developed for the detection of peste-des-petits-ruminants virus antigens in infected caprine tissue homogenates. Dots of tissue homogenates were applied to nitrocellulose papers, and any unreacted sites were blocked with 5% skim milk powder in triethanolamine-buffered saline. After incubation of the papers in tissue culture supernatant monoclonal antibody against the peste-des-petits-ruminants virus, the antigen-antibody reaction was detected with peroxidase-conjugated anti-mouse immunoglobulin G and the enzyme substrate 4-chloro-1-naphthol. Positive results were visualized as blue dots. Results of the dot enzyme immunoassay compared favorably with those of the standard enzyme-linked immunosorbent assay. Incorporation of Nonidet P-40 in the washing solution did not improve the sensitivity of the dot enzyme immunoassay, and pretreatment of homogenates with Nonidet P-40 before application to the nitrocellulose paper inhibited the binding of the antigen to the paper and reduced the intensity of the color development.