Development and evaluation of ELISA procedures to detect antibodies against the major envelope protein (G(L)) of equine arteritis virus.
Enzyme-linked immunosorbant assays (ELISAs) were developed for the detection of antibodies against the major envelope glycoprotein (G(L)) of equine arteritis virus (EAV). A 6-Histidine tagged recombinant protein expressing the complete G(L) ectodomain (G(L)-6His), a glutathione-S-transferase recombinant protein expressing amino acids 55-98 of G(L) (G(L)-GST) and an ovalbumin-conjugated synthetic peptide representing amino acids 81-106 of G(L) (G(L)-OVA) were used as diagnostic antigens. An ELISA procedure was developed and optimised for each antigen. The G(L)-OVA and G(L)-6His assays showed the greatest specificity while the G(L)-GST assay was slightly more sensitive that the G(L)-OVA and G(L)-6His assays; results based on the analysis of 50 virus neutralisation positive and 50 virus neutralisation negative sera. The G(L)-OVA ELISA was selected for further evaluation since it was simpler to use than ELISAs based on recombinant antigens and did not suffer from background reactivity. The final sensitivity and specificity of the G(L)-OVA ELISA were 96.75 and 95.6%, respectively, results based on the analysis of 400 virus neutralisation positive and 400 virus neutralisation negative sera. It also detected EAV antibody (100% efficiency) in seropositive shedding stallions and, in ponies infected experimentally with the UK93 isolate of EAV, the appearance of virus neutralising antibodies and G(L)-OVA ELISA-specific immunoglobulins coincided.
Nugent
J
J
Centre for Preventive Medicine, Animal Health Trust, Lanwades Park, Kentford, Newmarket, CB8 7UU, Suffolk, UK. josie.nugent@aht.org.uk
Sinclair
R
R
deVries
A A
AA
Eberhardt
R Y
RY
Castillo-Olivares
J
J
Davis Poynter
N
N
Rottier
P J
PJ
Mumford
J A
JA
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis* / Q000097:blood; D000956:Antigens, Viral / Q000276:immunology; D018174:Arterivirus Infections / Q000662:veterinary*; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary*; D007240:Equartevirus / Q000276:immunology*; D005982:Glutathione Transferase / Q000276:immunology / Q000378:metabolism; D006639:Histidine / Q000276:immunology; D006734:Horse Diseases / Q000175:diagnosis* / Q000276:immunology; D006736:Horses; D010047:Ovalbumin / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012680:Sensitivity and Specificity; D006113:United Kingdom; D014759:Viral Envelope Proteins / Q000276:immunology*
D000914:Antibodies, Viral; D000956:Antigens, Viral; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C105880:glycoprotein L, equine arteritis virus; D006639:Histidine; D010047:Ovalbumin; D005982:Glutathione Transferase