An ELISA for detection of antibodies against porcine epidemic diarrhoea virus (PEDV) based on the specific solubility of the viral surface glycoprotein
Knuchel, M.; Ackermann, M.; Müller, H.K.; Kihm, U.
Veterinary Microbiology 32(2): 117-134
Viral proteins of porcine epidemic diarrhoea virus (PEDV) were extracted from the cytoplasm of infected Vero cells using hypotonic conditions and a non-ionic detergent. Both the pH and the NaCl concentration of the extraction buffer were varied in attempts to increase the solubility of the virion spike glycoproteins (S-protein) and of the nucleocapsid proteins (N-protein). Monoclonal antibodies, hyperimmune sera and convalescent pig sera were used to identify and monitor these proteins by immunoprecipitation and Western blots. The solubility of the S-protein was optimal at pH 4, whereas that of the N-protein was optimal at pH 9. Consequently, it was possible to enrich for either S-protein or N-protein; increases in the NaCl concentration of the buffer were of no advantage in this respect. Enriched preparations of the S-protein and N-protein were used as ELISA antigen for the S-ELISA and N-ELISA, respectively. The S-ELISA proved to be the more effective of the two immunoassays. Antibodies against S-protein remained detectable for longer periods of time than anti-N-protein antibodies in the sera of PEDV-infected pigs. Using this ELISA of increased sensitivity, it was observed that only a small number of farms in Switzerland had been infected with PEDV.