Detection of antibodies to caprine arthritis-encephalitis virus using recombinant gag proteins.
The coding sequences of the core proteins p17 and p28 of caprine arthritis-encephalitis virus (CAEV) were amplified using the polymerase chain reaction and cloned into the plasmid expression vector p-GEX-2T. Both p17 and p28 were expressed as fusion proteins with glutathione S-transferase. The recombinant proteins were affinity purified from induced bacterial lysates using glutathione-agarose beads. The purified proteins were used in an enzyme-linked immunosorbent assay (ELISA) to detect antibodies against CAEV in goat sera and milk samples. Three different ELISA tests were developed based on p17, p28 or the combination of these two recombinant proteins (p17 + p28). A comparison was made to an ELISA based on purified whole virus particles and to agar immunodiffusion test (AGID). Sera with conflicting results in the different ELISA tests were examined by Western blotting. There was a high correlation between the ELISA tests based on p17 + p28 recombinant proteins and whole virus ELISA, with an estimated kappa value of 0.92. Only 72-75% of the sera that tested positive in these two ELISA tests were positive in AGID. Antibodies to CAEV were detected in significantly more animals when serum samples were tested compared to milk samples. Based on the time and materials required to prepare the reagents, the recombinant based ELISA test was less expensive than the whole virus ELISA.
Rimstad
E
E
Department of Medicine, School of Veterinary Medicine, University of California, Davis.
East
N
N
DeRock
E
E
Higgins
J
J
Pedersen
N C
NC
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis* / Q000097:blood; D016088:Arthritis-Encephalitis Virus, Caprine / Q000276:immunology*; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary; D004926:Escherichia coli; D015683:Gene Products, gag / Q000276:immunology* / Q000302:isolation & purification; D015511:Goat Diseases / Q000276:immunology / Q000382:microbiology; D006041:Goats; D016180:Lentivirus Infections / Q000276:immunology / Q000382:microbiology / Q000662:veterinary; D008892:Milk / Q000382:microbiology; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000662:veterinary; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000302:isolation & purification; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral; D017931:DNA Primers; D015683:Gene Products, gag; D011993:Recombinant Fusion Proteins