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- Title
Optimization of a Non-Radioactive High-Throughput Assay for Decarboxylase Enzymes.
- Authors
David C. Smithson; Anang A. Shelat; Jeffrey Baldwin; Margaret A. Phillips; R. Kiplin Guy
- Abstract
Herein, we describe the optimization of a linked enzyme assay suitable for high-throughput screening of decarboxylases, a target family whose activity has historically been difficult to quantify. Our approach uses a commercially available bicarbonate detection reagent to measure decarboxylase activity. The assay is performed in a fully enclosed automated screening system under inert nitrogen atmosphere to minimize perturbation by exogenous CO2. Receiver operating characteristic (ROC) analysis following a pilot screen of a small library of ∼3,600 unique molecules for inhibitors ofTrypanosoma brucei ornithine decarboxylase quantitatively demonstrates that the assay has excellent discriminatory power (area under the curve 0.90 with 95% confidence interval between 0.82 and 0.97).
- Subjects
MATHEMATICAL optimization; HIGH throughput screening (Drug development); DECARBOXYLASES; ENZYME-linked immunosorbent assay; TARGETED drug delivery; CHEMICAL reagents; TRYPANOSOMA brucei; ENZYME inhibitors
- Publication
Assay & Drug Development Technologies, 2010, Vol 8, Issue 2, p175
- ISSN
1540-658X
- Publication type
Article
- DOI
10.1089/adt.2009.0249