NDLI: Confocal Nanoscanning, Bead Picking (CONA): PickoScreen Microscopes for Automated and Quantitative Screening of One-Bead One-Compound Libraries

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Hintersteiner, Martin Buehler, Christof Uhl, Volker Schmied, Mario Müller, Jürgen Kottig, Karsten Auer, Manfred
Description	Country affiliation: United kingdom Author Affiliation: Hintersteiner M ( School of Biological Sciences, The University of Edinburgh, Michael Swann Building, 3.34, The King's Buildings, Mayfield Road, Edinburgh, EH9 3JR, UK.)
Abstract	Solid phase combinatorial chemistry provides fast and cost-effective access to large bead based libraries with compound numbers easily exceeding tens of thousands of compounds. Incubating one-bead one-compound library beads with fluorescently labeled target proteins and identifying and isolating the beads which contain a bound target protein, potentially represents one of the most powerful generic primary high throughput screening formats. On-bead screening (OBS) based on this detection principle can be carried out with limited automation. Often hit bead detection, i.e. recognizing beads with a fluorescently labeled protein bound to the compound on the bead, relies on eye-inspection under a wide-field microscope. Using low resolution detection techniques, the identification of hit beads and their ranking is limited by a low fluorescence signal intensity and varying levels of the library beads’ autofluorescence. To exploit the full potential of an OBS process, reliable methods for both automated quantitative detection of hit beads and their subsequent isolation are needed. In a joint collaborative effort with Evotec Technologies (now Perkin-Elmer Cellular Technologies Germany GmbH), we have built two confocal bead scanner and picker platforms PS02 and a high-speed variant PS04 dedicated to automated high resolution OBS. The PS0X instruments combine fully automated confocal large area scanning of a bead monolayer at the bottom of standard MTP plates with semiautomated isolation of individual hit beads via hydraulic-driven picker capillaries. The quantification of fluorescence intensities with high spatial resolution in the equatorial plane of each bead allows for a reliable discrimination between entirely bright autofluorescent beads and real hit beads which exhibit an increased fluorescence signal at the outer few micrometers of the bead. The achieved screening speed of up to 200 000 bead assayed in less than 7 h and the picking time of ∼1 bead/min allow exploitation of one-bead one-compound libraries with high sensitivity, accuracy, and speed.
File Format	HTM / HTML
ISSN	21568952
Issue Number	5
Volume Number	11
e-ISSN	21568944
Journal	Journal of Combinatorial Chemistry
Language	English
Publisher	American Chemical Society
Publisher Date	2009-09-01
Publisher Place	United States
Access Restriction	Subscribed
Subject Keyword	Discipline Chemistry Automation Combinatorial Chemistry Techniques Microscopy, Confocal Instrumentation Algorithms Fluorescent Dyes Chemistry Methods Proteins Journal Article
Content Type	Text
Resource Type	Article
Subject	Chemistry Medicine

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