AlphaScreen-Based Assays: Ultra-High-Throughput Screening for Small-Molecule Inhibitors of Challenging Enzymes and Protein-Protein Interactions

Methods Mol Biol. 2016:1439:77-98. doi: 10.1007/978-1-4939-3673-1_5.

Abstract

AlphaScreen technology has been routinely utilized in high-throughput screening assays to quantify analyte accumulation or depletion, bimolecular interactions, and post-translational modifications. The high signal-to-background, dynamic range, and sensitivity associated with AlphaScreens as well as the homogenous assay format and reagent stability make the technology particularly well suited for high-throughput screening applications. Here, we describe the development of AlphaScreen assays to identify small-molecule inhibitors of enzymes and protein-protein interactions using the highly miniaturized 1536-well format. The subsequent implementation of counter assays to identify false-positive compounds is also discussed.

Keywords: AlphaLISA; AlphaScreen; Assay; ELISA; Enzyme; HTS; High-throughput screening; Protein-protein interaction.

Publication types

  • Research Support, N.I.H., Intramural

MeSH terms

  • Animals
  • Cell Line
  • Drug Evaluation, Preclinical / methods*
  • Enzyme Assays / methods*
  • Enzyme Inhibitors / pharmacology*
  • High-Throughput Screening Assays / methods*
  • Humans
  • Luminescent Measurements / methods*
  • Protein Interaction Maps / drug effects*
  • Small Molecule Libraries / pharmacology*

Substances

  • Enzyme Inhibitors
  • Small Molecule Libraries