[Improved test for the detection of reverse transcriptase of bovine leukosis virus]

Abstract

The revertase test with exogenous matrix, poly-(rA) . oligo-(dT), for the detection of bovine leukemia virus from crude virus sediments was standardised and miniaturised. An amount of 10 ml of cell culture supernatant of short-term cultured lymphocytes (5 . 10(6) cells/ml) is quite sufficient for testing one cattle sample. The lower sensitivity limit of the test was found to be 10(6) virus particles. The test is properly reproducible, within tolerance limits of +/- 20--30 per cent, provided that optimum lysis conditions be maintained (0.01 per cent triton X-100, 20 minutes, 0--4 degrees C incubation) and under the condition that the protein quantity in 100 microliter test solution does not exceed the threshold of 3--15 micrograms. The specificity of the test is based on the use of free viruses from cell culture supernatant, the optimum temperature of the revertase reaction at 25 degrees C, which actually deviates from that for cellular DNA polymerases, that is 37 degrees C, and magnesium ion concentration which has to be optimum for bovine leucosis virus revertase. Two-hundred heads of cattle, differing by haematological status, were examined, and 56 per cent of them were, clearly, virus producers, among them 95 per cent of all animals with positively established leucosis and 36 per cent of the haematologically intact animals. Examinations of individuals have shown that in repetitive checks, carried out in intervals between two months and one week, the revertase activities varied by something between 0.5 and two magnitudes.