An ELISA for detection of antibodies against porcine epidemic diarrhoea virus (PEDV) based on the specific solubility of the viral surface glycoprotein.
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.
Knuchel
M
M
Institut für Virologie, Vet.-med. Fakultät, Universität Zürich, Switzerland.
Ackermann
M
M
Müller
H K
HK
Kihm
U
U
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000737:chemistry / Q000276:immunology; D002213:Capsid / Q000737:chemistry / Q000276:immunology; D003332:Coronaviridae / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000737:chemistry / Q000276:immunology*; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D008970:Molecular Weight; D012995:Solubility; D013552:Swine; D014709:Vero Cells; D014758:Viral Core Proteins / Q000737:chemistry / Q000276:immunology; D014764:Viral Proteins / Q000737:chemistry / Q000276:immunology*
D000914:Antibodies, Viral; D000956:Antigens, Viral; D006023:Glycoproteins; D014758:Viral Core Proteins; D014764:Viral Proteins; C064484:spike protein S, Transmissible gastroenteritis virus