Menon Veena, Ranganathn Arun, Jorgensen Vincent H, Sabio Michael, Christoffersen Claus T, Uberti Michelle A, Jones Kenneth A, Babu Poda Suresh
High-Throughput Screening, Biological Research, Lundbeck Research USA, Inc. 215 College Road, Paramus, NJ 07652, USA.
Assay Drug Dev Technol. 2008 Dec;6(6):787-93. doi: 10.1089/adt.2008.157.
A luminescence assay using a new plate reader, the LumiLux (PerkinElmer, Waltham, MA), has been validated for high-throughput screening (HTS). In this study, we compared the aequorin luminescence-based calcium mobilization assay to the fluorescence-based calcium assay. A cell line stably co-expressing apo-aequorin, a chimeric G-protein, and a G-protein-coupled dopamine receptor was used to screen a collection of 8,106 compounds using the Hamamatsu Photonics (Bridgewater, NJ) FDSS6000 and LumiLux as the plate readers. The assay parameters evaluated included hit rate correlation, signal-to-noise ratio, and overall assay performance calculated by Z and standard deviation. The average Z values and hit rates were comparable between assay platforms;however, the standard deviation for the agonist aequorin assay was significantly smaller. There was also a significant decrease in the number of false-positives with the aequorin assay. These results suggest that the aequorin assay in combination with the new plate reader, LumiLux, provides a simple, cost-effective, robust, and sensitive assay for HTS
一种使用新型酶标仪LumiLux（珀金埃尔默公司，马萨诸塞州沃尔瑟姆）的发光测定法已被验证可用于高通量筛选（HTS）。在本研究中，我们将基于水母发光蛋白发光的钙动员测定法与基于荧光的钙测定法进行了比较。使用稳定共表达脱辅基水母发光蛋白、嵌合G蛋白和G蛋白偶联多巴胺受体的细胞系，以滨松光子学公司（新泽西州布里奇沃特）的FDSS6000和LumiLux作为酶标仪，对8106种化合物进行筛选。评估的测定参数包括命中率相关性、信噪比以及通过Z值和标准差计算的整体测定性能。各测定平台之间的平均Z值和命中率相当；然而，激动剂水母发光蛋白测定法的标准差显著更小。水母发光蛋白测定法的假阳性数量也显著减少。这些结果表明，水母发光蛋白测定法与新型酶标仪LumiLux相结合，为高通量筛选提供了一种简单、经济高效、稳健且灵敏的测定方法。
