Details

Autor(en) / Beteiligte

• Taura, Jaume
• López‐Cano, Marc
• Fernández‐Dueñas, Víctor
• Ciruela, Francisco

Titel

Visualizing G Protein‐Coupled Receptor‐Receptor Interactions in Brain Using Proximity Ligation In Situ Assay

Ist Teil von

• Current protocols, 2023-06, Vol.3 (6), p.e794-n/a

Ort / Verlag

United States

Erscheinungsjahr

2023

Quelle

Alma/SFX Local Collection

Beschreibungen/Notizen

• G protein‐coupled receptors (GPCRs) constitute the largest family of plasma membrane receptors and the main drug targets in therapeutics. GPCRs can establish direct receptor‐receptor interactions (oligomerization), which can also be considered as targets for drug development (GPCR oligomer‐based drugs). However, prior to designing any novel GPCR oligomer‐based drug development program, demonstrating the existence of a named GPCR oligomer in native tissues is needed as part of its target engagement definition. Here, we discuss the proximity ligation in situ assay (P‐LISA), an experimental approach that reveals GPCR oligomerization in native tissues. We provide a detailed step‐by‐step protocol to perform P‐LISA experiments and visualize GPCR oligomers in brain slices. We also provide instructions for slide observation, data acquisition, and quantification. Finally, we discuss the critical aspects determining the success of the technique, namely the fixation process and the validation of the primary antibodies used. Overall, this protocol may be used to straightforwardly visualize GPCR oligomers in the brain. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Visualization of GPCR oligomers by proximity ligation in situ assay (P‐LISA) Support Protocol: Slide observation, image acquisition, and quantification