Development of an intracellular quantitative assay to measure compound binding kinetics

Lay, Charles S.; Thomas, Daniel; Evans, John P.; Campbell, Matthew; McCombe, Kristopher D.; Phillipou, Alexander; Gordon, Laurie J.; Jones, Emma; Riching, Kristin M.; Mahmood, Mahnoor; Messenger, Cassie; Carver, Charlotte E.; Gatfield, Kelly M; Craggs, Peter D.

doi:10.1016/j.chembiol.2021.07.018

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Cited by 3 publications

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“…Fitting the BRET data allows measurement of forward and reverse binding rates and the determination of K D . For complete details on the use and execution of this profile, please refer to Lay et al (2021) .…”

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confidence: 99%

A kinetic intra-cellular assay (KICA) to measure quantitative compound binding kinetics within living cells

et al. 2022

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Summary The Kinetic Intra-Cellular Assay (KICA) is a recombinant cell-based technique that utilizes NanoBRET technology. KICA enables the measurement of intracellular binding kinetics. This protocol describes steps for cellular transfection and expression, followed by addition of a target specific fluorophore conjugated probe and a range of concentrations of competitor compounds, followed by the measurement of BRET in a 384 well format. Fitting the BRET data allows measurement of forward and reverse binding rates and the determination of K D . For complete details on the use and execution of this profile, please refer to Lay et al. (2021) .

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confidence: 99%