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CRAF-14-3-3 Interaction Analysis in Live Cells Using a BRET-Based Approach

CRAF-14-3-3 Interaction Analysis in Live Cells Using a BRET-Based Approach

March 01, 2024

Author Spotlight: Integrating BRET-Based Assays and Rare Mutation Analysis to Decipher RAF Kinase Regulation in Live Cells

01:07 min
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Transfection of 293FT Cells with pCMV5-NanoLuc-CRAF and pCMV5-14-3-3ζ-Halo Expression Constructs

02:01 min
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21 Views

Preparing 293FT Cells Transfected with pCMV5-NanoLuc-CRAF and pCMV5-14-3-3ζ-Halo for BRET Emission Assay

03:39 min
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27 Views
1Department of Cell and Molecular Pharmacology and Experimental Therapeutics,Medical University of South Carolina, 2Hollings Cancer Center,Medical University of South Carolina, 3Laboratory of Cell and Developmental Signaling, Center for Cancer Research,National Cancer Institute-Frederick

概要

This protocol describes a BRET-based assay for measuring the interactions of the CRAF kinase with 14-3-3 proteins in live cells. The protocol outlines steps for preparing the cells, reading BRET emissions, and data analysis. An example result with identification of appropriate controls and troubleshooting for assay optimization is also presented.

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