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Updated: Oct 6, 2025
Adhesion Frequency Assay for In Situ Kinetics Analysis of Cross-Junctional Molecular Interactions at the Cell-Cell Interface
Published on: November 2, 2011
A kinetic intra-cellular assay (KICA) to measure quantitative compound binding kinetics within living cells
Charles S Lay1,2,3, Daniel A Thomas3,4, John P Evans3
1Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham, Nottingham NG7 2UH, UK.
Abstract:
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 KD. For complete details on the use and execution of this profile, please refer to Lay et al. (2021).
