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Applications of titration-based compound sets for use in rapid evaluation of high-throughput screening assays

Zhang, Ji, Auld, Douglas, Hill, William, Ardayfio, Ophelia, Kang, Zhao Bin, Smith, Thomas, Wallace, Iain and Bowes, Scott (2013) Applications of titration-based compound sets for use in rapid evaluation of high-throughput screening assays. Journal Biomolecular Screening, 19 (5). pp. 651-660. ISSN 1087-05711552-454X

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: Pilot testing of an assay intended for high-throughput screening (HTS) with small compound sets is a necessary but often time consuming step in the validation of an assay protocol. Typically, assay performance statistical factors such as Zʹ-factor, repeatability, and hit rates as well as the automated protocol and logistics are assessed at a single concentration in replicate runs, followed by confirmation and validation of the hits at multiple concentrations to determine concentration-response curves. When the initial testing compound concentration is less than optimal, this can involve iterative testing at different concentrations to further evaluate hit rates, which can be time consuming. Quantitative HTS (qHTS) enables flexible and rapid collection of assay performance statistics, hits, and concentration-response curves against chemical libraries in a single experiment. Here we investigated the application of qHTS in pilot testing of HTS assays using selected sets of 1,400 to 5,000 compounds and show several advantages where this approach can facilitate pilot testing. We describe the process for producing eight point concentration response curves using an inter-plate asymmetric dilution protocol where the first four concentrations are used to represent the range of typical single concentrations employed in HTS hit identification and the last four concentrations are added for facilitating robust curve fitting and determining the potency/efficacy of the hits. We also describe how this data can be analyzed to predict the frequency of false positives, false negatives, hit rates, as well as confirmation rates for the HTS process as a function of screening concentration. By taking into account the compound pharmacology in the assay, this paradigm can enable rapid assessment of assay performance factors and choosing the optimal concentration for the large-scale HTS in one experiment.

Item Type: Article
Date Deposited: 13 Oct 2015 13:13
Last Modified: 04 Jul 2016 23:46


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