Protocol abstract
Nature Protocols 1, - 2344 - 2352 (2006)
Published online: 21 December 2006 | doi:10.1038/nprot.2006.282
Subject Categories: Biochemistry and protein analysis | Chemical modification | Imaging | Pharmacology and toxicology | Synthetic chemistry
A method for the covalent capture and screening of diverse small molecules in a microarray format
James E Bradner1,2, Olivia M McPherson1 & Angela N Koehler1
Abstract
This protocol describes a robust method for the covalent capture of small molecules with diverse reactive functional groups in microarray format, and outlines a procedure for probing small-molecule microarrays (SMMs) with proteins of interest. A vapor-catalyzed, isocyanate-mediated surface immobilization scheme is used to attach bioactive small molecules, natural products and small molecules derived from diversity-oriented synthesis pathways. Additionally, an optimized methodology for screening SMMs with purified proteins and cellular lysates is described. Finally, a suggested model for data analysis that is compatible with commercially available software is provided. These procedures enable a platform capability for discovering novel interactions with potential applications to immunoglobulin profiling, comparative analysis of cellular states and ligand discovery. With the appropriate materials and experimental setup, the printing of SMMs can be completed in 14 hours over 3 days. Screening and data analysis requires 2 days. A detailed timeline is provided.
- Broad Institute of Harvard and MIT, 7 Cambridge Center, Cambridge, Massachusetts 02142, USA.
- Division of Hematologic Neoplasia, Dana-Farber Cancer Institute, Harvard Medical School, 44 Binney Street, Boston, Massachusetts 02115, USA.
Correspondence to: Angela N Koehler1 e-mail: koehler@fas.harvard.edu
nature-products
A-Z product listing
- (Benzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate(Novabiochem)
- 1,6-diisocyanatohexane(Aldrich)
- 2-slide centrifuge for microarray drying(Sunergia Medical)
- 293T Cells(ATCC)
- 384-well polypropylene natural microarray plates, cylindrical wells(Abgene)
- 4-well rectangular polystyrene dishes(Nunc)
MORE ARTICLES LIKE THIS
These links to content published by NPG are automatically generated.
