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Related Experiment Video

Updated: Mar 9, 2026

Detection and Visualization of DNA Damage-induced Protein Complexes in Suspension Cell Cultures Using the Proximity Ligation Assay

Detection and Visualization of DNA Damage-induced Protein Complexes in Suspension Cell Cultures Using the Proximity Ligation Assay

Published on: June 9, 2017

Proximity Ligation Assay (PLA) to Evaluate DISC and MISC Composition.

Melissa Thomas^1,2,3, Patrick Legembre^4,5,6

¹Centre Eugène Marquis, rue Bataille Flandres Dunkerque, 35042, Rennes, France.

Methods in Molecular Biology (Clifton, N.J.)

|January 13, 2017

Summary

Identifying protein-protein interactions (PPIs) is crucial for understanding cell signaling. Proximity ligation assay (PLA) allows for specific, in situ detection and quantification of PPIs, like CD95/FADD interactions, within cells.

Keywords:

CD95 Death inducing signaling complex FADD Protein–protein interaction Proximity ligation assay

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Last Updated: Mar 9, 2026

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Published on: January 20, 2015

Area of Science:

Cell Biology
Molecular Biology
Immunology

Background:

The death receptor CD95 (also known as Fas or APO-1) plays a critical role in apoptosis, with its localization to lipid rafts and internalization influencing signaling pathways.
CD95 interacts with CD95 Ligand (CD95L) to form dynamic protein complexes, and understanding these protein-protein interactions (PPIs) is vital for predicting cellular responses.
Dysregulation of PPIs, including their subcellular distribution and intensity, is implicated in various diseases, necessitating precise methods for their evaluation.

Purpose of the Study:

To describe the application of the in situ proximity ligation assay (PLA) for identifying and localizing PPIs.
To validate the use of PLA in confirming the interaction between CD95 and FADD (Fas-Associated Death Domain protein).
To present PLA as a versatile method for discovering other CD95-interacting partners.

Main Methods:

In situ proximity ligation assay (PLA) was employed to detect and visualize PPIs within cells.
PLA allows for the subcellular localization of identified PPIs.
The method provides quantitative data on PPIs at the cellular level with high specificity, surpassing traditional immunofluorescence techniques.

Main Results:

The study successfully utilized PLA to confirm the interaction between CD95 and FADD.
The results demonstrate PLA's capability to pinpoint the subcellular location of PPIs.
PLA offers a specific and quantifiable approach to studying PPIs in cellular contexts.

Conclusions:

In situ proximity ligation assay (PLA) is a powerful technique for identifying and quantifying PPIs within their native subcellular environment.
PLA provides a highly specific alternative to conventional methods for studying protein interactions.
This protocol, demonstrated with CD95/FADD, can be broadly applied to investigate other CD95-mediated PPIs and advance our understanding of signaling pathways and disease pathogenesis.