After comparison of various homogenization buffers and clarification methods, tulip breaking virus (TBV) was purified on a large scale by homogenization in 0.1 M tris buffer (pH 9.0), clarification with Triton X-100, and differential centrifugation. The TBV suspensions were absorbed with an anti-host protein antiserum. The purity of the product was verified by mechanical inoculation of test plants, serology, electron microscopy, spectrophotometry, and analytical ultracentrifugation.The TBV antisera obtained from rabbits after intramuscular injections had titres of 1/1280 to 1/10240 in the microprecipitin test and were mainly used in conjugate dilutions of 1/1000 in ELISA. The absorbance values for healthy tulips and lilies were (very) low. The antisera prepared against TBV from tulip cv. Jack Laan reacted best with TBV in lilies, whereas the antisera prepared against TBV from tulip cv. Texas Flame reacted best with TBV in tulips.
Lily symptomless virus (LSV) was purified by clarification with chloroform, precipitation with 9 polyethylene glycol and NaC1, and differential centrifugation. The influence of the source material and some buffers on virus yields were determined.Antisera were prepared against intact and pyrrolidine degraded LSV. It was concluded that intact and degraded LSV have very few antigenic determinants in common or none at all. The sensitivities of the micro-precipitin test and the single immunodiffusion drop test were about the same, but lower than that of electron microscopy.In the testing of lilies for LSV the most reliable results in leaves were obtained during the period from two weeks after flowering until close to the end of the growing season, and in leaves growing at a level about one-fourth of the distance from the top of the stem. In contrast to secondary infections, primary infections were detected more successfully in stored bulbs than in leaves taken from plants in the preceding growing season.In the testing of tulips, LSV was detected better in flowers than in leaves. Detection of primary infections was almost impossible. Except for those with a pink flower, experimentally infected tulips remained symptomless.
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