Volume 91, Issue 1 e86
PROTOCOL

Detection of G Protein-Coupled Receptor Complexes in Postmortem Human Brain by Proximity Ligation Assay

Ying Zhu

Ying Zhu

Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York

Department of Psychiatry, Columbia University, New York, New York

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Andrew J. Dwork

Andrew J. Dwork

Department of Psychiatry, Columbia University, New York, New York

Department of Pathology and Cell Biology, Columbia University, New York, New York

Division of Molecular Imaging and Neuropathology, New York State Psychiatric Institute, New York, New York

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Pierre Trifilieff

Pierre Trifilieff

Univ. Bordeaux, INRAE, Bordeaux INP, NutriNeuro, UMR 1286, F-33000, Bordeaux, France

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Jonathan A. Javitch

Corresponding Author

Jonathan A. Javitch

Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York

Department of Psychiatry, Columbia University, New York, New York

Department of Pharmacology, Columbia University, New York, New York

Corresponding author: [email protected]Search for more papers by this author
First published: 13 January 2020
Citations: 3

Abstract

Combining immunological and molecular biological methods, the antibody-based proximity ligation assay (PLA) has been used for more than a decade to detect and quantify protein-protein interactions, protein modification, and protein expression in situ, including in brain tissue. However, the transfer of this technology to human brain samples requires a number of precautions due to the nature of the specimens and their specific processing. Here, we used the PLA brightfield detection technique to assess the expression of dopamine D2 receptor and adenosine A2A receptor and their proximity in human postmortem brains, and we developed a systematic random sampling method to help quantify the PLA signals. © 2019 by John Wiley & Sons, Inc.

Basic Protocol 1: Sample preparation and sectioning for PLA_BF

Basic Protocol 2: PLA_BF staining of brain tissue

Basic Protocol 3: Image acquisition and result analysis

Support Protocol: Luxol fast blue/cresyl violet staining