ArticleBiomolecular detection and quantification2015
Development, validation and quantitative assessment of an enzymatic assay suitable for small molecule screening and profiling: A case-study.
Article in Biomolecular detection and quantification, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 3 papers.
What it found
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
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Who cites it
3 citing papers in PubMed.
- Best Practices for Development and Validation of Enzymatic Activity Assays to Support Drug Development for Inborn Errors of Metabolism and Biomarker Assessment.The AAPS journal · 2024Review
- Optimization and Validation of an In Vitro Standardized Glycogen Phosphorylase Activity Assay.Molecules (Basel, Switzerland) · 2021Article
- Physiologically relevant orthogonal assays for the discovery of small-molecule modulators of WIP1 phosphatase in high-throughput screens.The Journal of biological chemistry · 2019Article
Corrections and comments
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Authors and funding
8 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The successful discovery and subsequent development of small molecule inhibitors of drug targets relies on the establishment of robust, cost-effective, quantitative, and physiologically relevant in vitro assays that can support prolonged screening and optimization campaigns. The current study illustrates the process of developing and validating an enzymatic assay for the discovery of small molecule inhibitors using alkaline phosphatase from bovine intestine as model target. The assay development workflow includes an initial phase of optimization of assay materials, reagents, and conditions, continues with a process of miniaturization and automation, and concludes with validation by quantitative measurement of assay performance and signal variability. The assay is further evaluated for dose-response and mechanism-of-action studies required to support structure-activity-relationship studies. Emphasis is placed on the most critical aspects of assay optimization and other relevant considerations, including the technology, assay materials, buffer constituents, reaction conditions, liquid handling equipment, analytical instrumentation, and quantitative assessments. Examples of bottlenecks encountered during assay development and strategies to address them are provided.
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