Detection of G Protein-Coupled Receptor Complexes in Postmortem Human Brain by Proximity Ligation Assay
Ying Zhu
Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York
Department of Psychiatry, Columbia University, New York, New York
Search for more papers by this authorAndrew 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
Search for more papers by this authorPierre Trifilieff
Univ. Bordeaux, INRAE, Bordeaux INP, NutriNeuro, UMR 1286, F-33000, Bordeaux, France
Search for more papers by this authorCorresponding 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 authorYing Zhu
Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York
Department of Psychiatry, Columbia University, New York, New York
Search for more papers by this authorAndrew 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
Search for more papers by this authorPierre Trifilieff
Univ. Bordeaux, INRAE, Bordeaux INP, NutriNeuro, UMR 1286, F-33000, Bordeaux, France
Search for more papers by this authorCorresponding 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 authorAbstract
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
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