Detection of G Protein‐Coupled Receptor Complexes in Postmortem Human Brain by Proximity Ligation Assay
Current Protocols in Neuroscience. 2020-01-13; 91(1):
DOI: 10.1002/cpns.86
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Zhu Y(1)(2), Dwork AJ(2)(3)(4), Trifilieff P(5), Javitch JA(1)(2)(6).
Author information:
(1)Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York.
(2)Department of Psychiatry, Columbia University, New York, New York.
(3)Department of Pathology and Cell Biology, Columbia University, New York, New York.
(4)Division of Molecular Imaging and Neuropathology, New York State Psychiatric Institute, New York, New York.
(5)Univ. Bordeaux, INRAE, Bordeaux INP, NutriNeuro, UMR 1286, F-33000, Bordeaux, France.
(6)Department of Pharmacology, Columbia University, New York, New York.
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.