A wide-field fluorescence microscope extension for ultrafast screening of one-bead one-compound libraries using a spectral image subtraction approach

Wolf Heusermann, Beat Ludin, Nhan T. Pham, Manfred Auer, Thomas Weidemann, Martin Hintersteiner*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

The increasing involvement of academic institutions and biotech companies in drug discovery calls for cost-effective methods to identify new bioactive molecules. Affinity-based on-bead screening of combinatorial one-bead one-compound libraries combines a split-mix synthesis design with a simple protein binding assay operating directly at the bead matrix. However, one bottleneck for academic scale on-bead screening is the unavailability of a cheap, automated, and robust screening platform that still provides a quantitative signal related to the amount of target protein binding to individual beads for hit bead ranking. Wide-field fluorescence microscopy has long been considered unsuitable due to significant broad spectrum autofluorescence of the library beads in conjunction with low detection sensitivity. Herein, we demonstrate how such a standard microscope equipped with LED-based excitation and a modern CMOS camera can be successfully used for selecting hit beads. We show that the autofluorescence issue can be overcome by an optical image subtraction approach that yields excellent signal-to-noise ratios for the detection of bead-associated target proteins. A polymer capillary attached to a semiautomated bead-picking device allows the operator to efficiently isolate individual hit beads in less than 20 s. The system can be used for ultrafast screening of >200,000 bead-bound compounds in 1.5 h, thereby making high-throughput screening accessible to a wider group within the scientific community.

Original languageEnglish
Pages (from-to)209-219
Number of pages12
JournalACS Combinatorial Science
Volume18
Issue number5
Early online date8 Apr 2016
DOIs
Publication statusPublished - 9 May 2016

Keywords

  • one-bead one-compound libraries
  • ultrafast screening
  • high-throughput screening
  • spectral imaging
  • COMBINATORIAL LIBRARIES
  • RAPID IDENTIFICATION
  • PEPTIDE LIBRARY
  • DRUG DISCOVERY
  • LIGANDS
  • DOMAINS
  • HITS

Fingerprint

Dive into the research topics of 'A wide-field fluorescence microscope extension for ultrafast screening of one-bead one-compound libraries using a spectral image subtraction approach'. Together they form a unique fingerprint.

Cite this