Acoustic Mist Ionization-Mass Spectrometry: A Comparison to Conventional High-Throughput Screening and Compound Profiling Platforms.

Drug discovery usually begins with a high-throughput screen (HTS) of thousands to millions of molecules to identify starting points for medicinal chemistry. Conventional HTS platforms require expensive reagents and typically have complex assay formats. HTS platforms based on radioactivity are expensive, both in terms of reagent cost and disposal. Furthermore, nonspecific interferences common to these technologies result in an extensive attrition of hits during validation experiments. Mass spectrometry (MS) is a highly selective, label-free technology that can quantify multiple analytes in a single experiment. However, most commercial MS platforms typically involve a separation or cleanup prior to analysis and are too slow for large-scale screening campaigns. Recently, an MS platform (AMI-MS) was introduced that uses acoustically generated droplets to deliver analyte molecules directly from microtiter plates into the mass spectrometer at subsecond per well sampling rates. Here, we demonstrate the application of AMI-MS by developing an HTS-compatible assay that measures the inhibition of histone acetyltransferase activity. Real-time kinetic measurements from a single well were used to determine enzyme Km and Vmax values. We compare the AMI-MS readout with conventional platforms in single-shot screening and multipoint profiling modes. The AMI-MS assay identified 86% of hits previously identified, with a pIC50 ≥ 5.0, in a scintillation proximity assay (SPA) HTS at a lower hit rate and with a significantly reduced cost per well compared to the SPA-based readout. Furthermore, pIC50s, as measured by AMI-MS, showed a good correlation with values generated by RapidFire-MS. AMI-MS has the potential to provide significant improvements to high-throughput bioassays.

Citation

Arseniy M Belov, Joseph Kozole, Mark F Bean, Carl A Machutta, Guofeng Zhang, Enoch N Gao, Luke Ghislain, Sammy S Datwani, Melanie Leveridge, Roland S Annan. Acoustic Mist Ionization-Mass Spectrometry: A Comparison to Conventional High-Throughput Screening and Compound Profiling Platforms. Analytical chemistry. 2020 Oct 20;92(20):13847-13854

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PMID: 32970424

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