Production of a baculovirus-derived gp50 protein and utilization in a competitive enzyme-linked immunosorbent assay for the serodiagnosis of pseudorabies virus.
The pseudorabies virus (PRV) gp50 envelope glycoprotein gene was cloned and expressed in a recombinant baculovirus. An anti-gp50 Mab (1842) recognized a protein of approximately 40 kDa in immunoblotting assays from infected insect cell lysates, while this product was not present in cells infected with wild-type baculovirus. The recombinant protein was purified by lectin affinity chromatography, utilizing lectins specific for O-linked oligosaccharides (Artocarpus integrifolia and Glycine max). Competitive (c) ELISAs, using either crude or lectin-purified antigen, were devised for the detection of antibodies to PRV in sera, and were capable of monitoring sero-conversion by day 14 post-infection. Furthermore, a specificity of 100% and sensitivity of 98% (crude lysate antigen) or 96% (lectin-purified antigen) was found for a panel of 80 swine sera, using the cELISA, as compared to a serum neutralization (SN) test. These studies demonstrated that recombinant PRV gp50 protein shows promise as a cELISA antigen, for serodetection of PRV.
Prud'homme
I
I
Animal Diseases Research Institute, Nepean, Ontario.
Zhou
E M
EM
Traykova
M
M
Trotter
H
H
Chan
M
M
Afshar
A
A
Harding
M J
MJ
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D000918:Antibody Specificity; D000956:Antigens, Viral / Q000276:immunology; D016367:Baculoviridae / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity / Q000379:methods / Q000662:veterinary; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods / Q000662:veterinary; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D005822:Genetic Vectors; D011558:Herpesvirus 1, Suid / Q000276:immunology*; D015151:Immunoblotting / Q000379:methods / Q000662:veterinary; D007668:Kidney / Q000166:cytology / Q000821:virology; D009036:Moths / Q000166:cytology / Q000821:virology; D009500:Neutralization Tests / Q000379:methods / Q000662:veterinary; D011557:Pseudorabies / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D011994:Recombinant Proteins / Q000378:metabolism; D012680:Sensitivity and Specificity; D013552:Swine; D013553:Swine Diseases / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000378:metabolism*
D000914:Antibodies, Viral; D000956:Antigens, Viral; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins