title: A rapid streptavidin-capture ELISA specific for the detection of antibodies to feline foamy virus.
abstract: We report a simple procedure for the rapid development of an ELISA with the potential for wide application to any defined protein antigen. The procedure involves the expression of protein encoded by a PCR product, using a commercially available T-vector that adds a biotin tag, and a single step purification by affinity for streptavidin for direct use in ELISA. In our experiments, a recombinant protein from the nucleocapsid domain of the feline foamy virus gag gene was expressed as a fusion protein with a biotin tag and then applied directly to streptavidin-coated ELISA wells. An extract from a clone with the insert in antisense orientation was used as a control. Non-specific reactions with antigen extracts from both sense and antisense clones were observed in 6 of the 376 (1.6%) sera tested. Antibody to feline foamy virus, which forms a stable persistent infection in cats, was detected in 107 of 201 (53%) Australian cats, but none of 175 sera from veterinarians. There was a 100% correlation between FeFV antibody detected by ELISA, immunoblot, serum neutralisation and virus isolation, confirming that this test is sensitive and specific.
authors.affiliation: School of Pharmacy and Medical Science, University of South Australia, Adelaide, Australia.
authors.forename: I G
authors.initials: IG
authors.lastname: Winkler
authors.forename: M
authors.initials: M
authors.lastname: Löchelt
authors.forename: J P
authors.initials: JP
authors.lastname: Levesque
authors.forename: J
authors.initials: J
authors.lastname: Bodem
authors.forename: R M
authors.initials: RM
authors.lastname: Flügel
authors.forename: R L
authors.initials: RL
authors.lastname: Flower
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000302:isolation & purification*; D000918:Antibody Specificity; D016375:Antisense Elements (Genetics); D002415:Cats; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D015870:Gene Expression; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D009500:Neutralization Tests; D016133:Polymerase Chain Reaction; D015995:Prevalence; D011994:Recombinant Proteins / Q000276:immunology; D011995:Recombination, Genetic; D012192:Retroviridae Infections / Q000453:epidemiology / Q000276:immunology; D016092:Spumavirus / Q000235:genetics / Q000254:growth & development / Q000276:immunology*; D019809:Streptavidin / Q000378:metabolism*; D018634:Veterinarians
chemical_list: D000914:Antibodies, Viral; D016375:Antisense Elements (Genetics); D004279:DNA, Viral; D015683:Gene Products, gag; D011994:Recombinant Proteins; D019809:Streptavidin