Simple and rapid method for production of whole-virus antigen for serodiagnosis of caprine arthritis-encephalitis virus by enzyme-linked immunosorbent assay

Citation
C. Simard et al., Simple and rapid method for production of whole-virus antigen for serodiagnosis of caprine arthritis-encephalitis virus by enzyme-linked immunosorbent assay, CL DIAG LAB, 8(2), 2001, pp. 352-356
Citations number
25
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Immunology
Journal title
CLINICAL AND DIAGNOSTIC LABORATORY IMMUNOLOGY
ISSN journal
1071-412X → ACNP
Volume
8
Issue
2
Year of publication
2001
Pages
352 - 356
Database
ISI
SICI code
1071-412X(200103)8:2<352:SARMFP>2.0.ZU;2-N
Abstract
Polyethylene glycol (PEG) was used to produce whole-virus antigen derived f rom tissue culture cells infected with a Canadian strain of caprine arthrit is-encephalitis virus. PEG antigen batches mere obtained after precipitatio n and concentration of infected tissue culture material with PEG 8000 and f inal treatment with sodium dodecyl sulfate. The optimum time of harvest of tissue culture extracted material to produce the maximum amount of viral pr oteins was determined in roller bottles, after cocultivation of infected an d noninfected fetal lamb corneal cells. Samples from day 9 to day 25 postcu lture were collected and processed. Py Western blotting, the optimum time o f harvest was found to be day 25 following the coculture. Two large batches of PEG antigen were prepared at the optimum time of harvest. Both batches gave similar results when tested by Western blotting and enzyme-linked immu nosorbent assay (ELISA), using reference control sera from infected and non infected goats. For further testing in ELISA, cutoff values and ratios were determined for PEG batch 1, using 200 known serum samples from goats free of the disease. The PEG antigen batch was compared with an in-house ELISA a ntigen in a kinetic mode, using 498 serum samples from field goats. The in- house ELISA antigen was produced following two rounds of ultracentrifugatio n and treatment with sodium dodecyl sulfate (R. A Heckert, W. B. McNab, S. M. Richardson, and M. R. Briscoe, Can. J. Vet. Res. 56:237-241, 1992). The PEG antigen batch was found suitable for ELISA, with a relative specificity of 100% and a relative sensitivity of 99.4% compared to the in-house ELISA antigen. This method of antigen production for ELISA was found to be rapid , inexpensive, and reliable for the diagnosis of caprine-arthritis encephal itis, without requiring the use of sophisticated laboratory equipment.