Lee M J, Goldsworthy G J
Department of Biology, Birkbeck College, University of London, England.
Anal Biochem. 1995 Jun 10;228(1):155-61. doi: 10.1006/abio.1995.1327.
A method is described for the preparation of metabolically active and hormone-sensitive dispersed cells from fat body of Locusta migratoria and their subsequent use in a filtration plate assay. This assay measures the uptake of radiolabelled acetate into the cells, and the use of dispersed cells reduces the time and labor involved in preparing dose-response curves to different test peptides and increases the precision of the estimate of potency. As a consequence of the reduced variability, fewer replicates per dose are required and, when operated as a plate assay, up to eight dose-response curves can be constructed per plate. The method described here for measuring metabolite uptake into cells and thus determining the potencies of adipokinetic hormones could be applied to the assay of both catabolic (glucagon-like) and anabolic (insulin-like) substances, not only in other insects, but also in mammalian cells. It could be used, therefore, as a suitable screening method for novel active compounds.
本文描述了一种从飞蝗脂肪体中制备具有代谢活性且对激素敏感的分散细胞的方法，以及这些细胞随后在滤板分析中的应用。该分析可测量放射性标记的乙酸盐在细胞中的摄取，使用分散细胞可减少制备不同测试肽的剂量反应曲线所需的时间和工作量，并提高效价估计的精度。由于变异性降低，每个剂量所需的重复次数减少，并且当作为板分析操作时，每块板最多可构建八条剂量反应曲线。这里描述的用于测量细胞对代谢物的摄取从而确定脂肪动激素效价的方法，不仅可应用于其他昆虫，还可应用于哺乳动物细胞中分解代谢（胰高血糖素样）和合成代谢（胰岛素样）物质的分析。因此，它可作为一种适用于新型活性化合物的筛选方法。
