Detection of G Protein‐Coupled Receptor Complexes in Postmortem Human Brain by Proximity Ligation Assay. Issue 1 (13th January 2020)

• Record Type: Journal Article
• Authors: Zhu, Ying; Dwork, Andrew J.; Trifilieff, Pierre; Javitch, Jonathan A.
• Editors: Crawley, Jacqueline N.; Gerfen, Charles R.; Rogawski, Michael A.; Sibley, David R.; Skolnick, Phil; Wray, Susan
• Is Part Of: Current protocols in neuroscience. Issue 91:Issue 1(2020)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2020-01-13
• Subjects: brightfield -- D2R‐A2AR complexes -- GPCR -- human brain -- PLA -- postmortem -- proximity ligation assay -- systematic random sampling; Neurosciences -- Laboratory manuals; Neurosciences; Cytological Techniques; Histological Techniques; Nervous System -- anatomy & histology; Neurosciences; Laboratory Manual; Laboratory manuals; 612.8
• DOI: 10.1002/cpns.86
• Languages: English
• ISSNs: 1934-8576

Abstract

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