Protocol to measure and analyze protein interactions in mammalian cells using bioluminescence resonance energy transfer.

DUVAL, Carla; STEFFEN, Candy; PAVIC, Karolina; ABANKWA, Daniel

doi:10.1016/j.xpro.2024.103348

Article (Scientific journals)

Protocol to measure and analyze protein interactions in mammalian cells using bioluminescence resonance energy transfer.

DUVAL, Carla; STEFFEN, Candy; PAVIC, Karolina et al.

2024 • In STAR Protocols, 5 (4), p. 103348

Peer Reviewed verified by ORBi

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https://hdl.handle.net/10993/62559

DOI
10.1016/j.xpro.2024.103348

PubMed
39342617

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Keywords :

Cell Biology; Cell-based Assays; Molecular Biology; Neuroscience (all); Biochemistry, Genetics and Molecular Biology (all); Immunology and Microbiology (all)

Abstract :

[en] Bioluminescence resonance energy transfer (BRET) allows to quantitate protein interactions in intact cells. Here, we present a protocol for measuring BRET due to transient interactions of oncogenic K-RasG12V in plasma membrane nanoclusters of HEK293-EBNA cells. We describe steps for seeding, transfecting, and replating cells. We then detail procedures for their preparation for BRET measurements on a CLARIOstar microplate reader and detailed data analysis. For complete details on the use and execution of this protocol, please refer to Steffen et al.1.

Disciplines :

Biochemistry, biophysics & molecular biology

Author, co-author :

DUVAL, Carla ; University of Luxembourg > Faculty of Science, Technology and Medicine (FSTM) > Department of Life Sciences and Medicine (DLSM)

STEFFEN, Candy ; University of Luxembourg > Faculty of Science, Technology and Medicine > Department of Life Sciences and Medicine > Team Daniel ABANKWA

PAVIC, Karolina ; University of Luxembourg > Faculty of Science, Technology and Medicine (FSTM) > Department of Life Sciences and Medicine (DLSM)

ABANKWA, Daniel ; University of Luxembourg > Faculty of Science, Technology and Medicine (FSTM) > Department of Life Sciences and Medicine (DLSM)

External co-authors :

Language :

English

Title :

Protocol to measure and analyze protein interactions in mammalian cells using bioluminescence resonance energy transfer.

Publication date :

28 September 2024

Journal title :

STAR Protocols

eISSN :

2666-1667

Publisher :

Cell Press, United States

Volume :

Issue :

Pages :

103348

Peer reviewed :

Peer Reviewed verified by ORBi

Additional URL :

https://api.elsevier.com/content/article/PII:S2666166724005136?httpAccept=text/xml

Funders :

National Research Fund

Funding text :

This work was supported by grants from the National Research Fund Luxembourg (FNR) INTER/UKRI/19/14174764-RAS-NANOME and INTER/NWO/19/14061736 - HRAS-PPi to D.K.A. and AFR/17927850/Duval C./KRuptor to C.J.D. We are grateful to Dr. Ganesh babu Manoharan for implementing the BRET technology in the Abankwa lab. The graphical abstract was created using BioRender.com .

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since 18 November 2024

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Bibliography

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