Open AccessQuantitative High‐Throughput Screening Using a Coincidence Reporter Biocircuit
Author(s) -
Schuck Brittany W.,
MacArthur Ryan,
Inglese James
Publication year - 2017
Publication title -
current protocols in neuroscience
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.307
H-Index - 40
eISSN - 1934-8576
pISSN - 1934-8584
DOI - 10.1002/cpns.27
Subject(s) - false positive paradox , high throughput screening , coincidence , throughput , reporter gene , computational biology , computer science , chemistry , combinatorial chemistry , biochemical engineering , nanotechnology , materials science , biology , medicine , biochemistry , engineering , telecommunications , artificial intelligence , gene , gene expression , alternative medicine , pathology , wireless
Abstract Reporter‐biased artifacts—i.e., compounds that interact directly with the reporter enzyme used in a high‐throughput screening (HTS) assay and not the biological process or pharmacology being interrogated—are now widely recognized to reduce the efficiency and quality of HTS used for chemical probe and therapeutic development. Furthermore, narrow or single‐concentration HTS perpetuates false negatives during primary screening campaigns. Titration‐based HTS, or quantitative HTS (qHTS), and coincidence reporter technology can be employed to reduce false negatives and false positives, respectively, thereby increasing the quality and efficiency of primary screening efforts, where the number of compounds investigated can range from tens of thousands to millions. The three protocols described here allow for generation of a coincidence reporter (CR) biocircuit to interrogate a biological or pharmacological question of interest, generation of a stable cell line expressing the CR biocircuit, and qHTS using the CR biocircuit to efficiently identify high‐quality biologically active small molecules. © 2017 by John Wiley & Sons, Inc.