Journal
JOURNAL OF BIOMOLECULAR SCREENING
Volume 10, Issue 6, Pages 599-605Publisher
SAGE PUBLICATIONS INC
DOI: 10.1177/1087057105276083
Keywords
high-throughput screening; prostaglandin E; synthase; enzyme immunoassay
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Prostaglandin E-2 synthase (mPGES-1), the enzyme which catalyzes the synthesis of PGE(2), is induced during the inflammatory response. For this reason, mPGES-1 could be a potential therapeutic target. A high-throughput screening assay was developed to identify potential inhibitors of mPGES-1. The assay consisted of a 30-s mPGES-1 enzymatic reaction followed by the detection of PGE(2) by enzyme immunoassay (EIA). The enzymatic reaction was performed in a batch mode because the instability of the substrate (10 min) limited the number of plates assayed within a working day. The detection of the product by EIA was performed on 3 instruments requiring 14 different steps for complete automation. The authors describe here the optimization and implementation of a 2-part assay on a Thermo CRS robotic system. More than 315,000 compounds were tested, and a hit rate of 0.84% was obtained for this assay. Although the entire assay required multiple steps, the assay was successfully miniaturized and automated for a high-throughput screening campaign.
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