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