LUMINESCENCE ASSAY FOR PHARMECEUTICAL SCREENING

光蛋白aequorin的基因已经被整合到几个 哺乳动物细胞系。因为这种蛋白质的发光反映了 局部的钙离子浓度，对于细胞的分析是非常有用的 被潜在药物激活或阻断的Ca++信号 治疗价值。一个主要的实验困难在于非常低的 发射的光的级别，因为只有一个光子被 每个Aequorin分子，我们计算出只有 每个细胞20000个分子。我们计划修改一个光子计数装置 我们建造了一台仪器来测量青蛙卵母细胞的发光。这个 将使用光子计数作为一种测定药物活性的方法 影响细胞钙代谢的物质。大量的化学品将被 对活动进行筛选，因此仪器的自动化程度将与 有可能。我们有一项困难的工作，那就是注射少量(20 至50微升)液体和细胞悬浮液。如果预赛 结果是阳性的，我们会考虑购买自动取样器带走 和乐器一起。设计中的另一个困难是 许多药物的粘性，甚至对不锈钢也是如此。 因为即使是先前实验中的微小残留物也可能影响 发光，所有可重复使用的部件和管子都必须制造 特氟龙~。

The gene for the photoprotein aequorin has been incorporated into several mammalian cell lines. Because the luminescence of this protein reflects the local Ca++ concentration, it is very useful for assaying cellular Ca++ signals that are activated, or blocked, by drugs of potential therapeutic value. A major experimental difficulty lies in the very low level of light that is emitted, since only one photon is liberated by each molecule of aequorin, and we have calculated that there are only 20,000 molecules per cell. We plan to modify a photon-counting instrument that we built to measure luminescence from frog oocytes. The photon count will be used as an assay to determine the activity of drugs that influence cell Ca++ metaboism. Large numbers of chemicals will be screened for activity, so the instrument will be automated as much as possible. We have the difficult job of injecting small quantities (20 to 50 microliters) of fluids and cell suspensions. If the preliminary results are positive, we will consider purchasing an autosampler to go along with the instrument. Another of the difficulties in the design is the ~sticky~ character of many of the drugs, even to stainless steel. Since even small residues from prior experiments could affect the luminescence, all of the reusable pieces and tubing will have to be made of Teflon~.