Optimizing In Situ Proximity Ligation Assays for Mitochondria, ER, or MERC Markers in Skeletal Muscle Tissue and Cells

Current protocols Pub Date : 2025-02-05 DOI:10.1002/cpz1.70043

Amber Crabtree, Han Le, Chanel Harris, Ashton Oliver, Andy Barillas, Johnathan Moore, Desiree Ngoc-ha Nguyen, Izabella Marie Rabago, Benjamin Rodriguez, Dominique C. Stephens, Heather K. Beasley, Edgar Garza-Lopez, Kit Neikirk, Margaret Mungai, Larry Vang, Zer Vue, Neng Vue, Andrea G. Marshall, Kyrin Turner, Jianqiang Shao, Sandra Murray, Jennifer A. Gaddy, Celestine Wanjalla, Jamaine Davis, Steven Damo, Antentor O. Hinton Jr

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Abstract

Proximity ligation assays (PLAs) use specific antibodies to detect endogenous protein-protein interactions. PLAs are a highly useful biochemical technique that allow two proteins within proximity to be visualized with fluorescent probes amplified by PCR. While this technique has gained prominence, the use of a PLA in mouse skeletal muscle (SkM) is novel. In this article, we discuss how the PLA method can be used in SkM to study the protein-protein interactions within mitochondria-endoplasmic reticulum contact sites (MERCs). © 2025 The Author(s). Current Protocols published by Wiley Periodicals LLC.

Basic Protocol: Proximity ligation assay for skeletal muscle tissue and myoblast for MERC proteins

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优化骨骼肌组织和细胞中线粒体、内质网或MERC标记物的原位接近结扎试验。

邻近连接法（pla）使用特异性抗体检测内源性蛋白-蛋白相互作用。pla是一种非常有用的生化技术，它允许用荧光探针通过PCR扩增看到两个接近的蛋白质。虽然这项技术已经获得了突出，但在小鼠骨骼肌（SkM）中使用聚乳酸是新颖的。在本文中，我们讨论了如何在SkM中使用PLA方法来研究线粒体-内质网接触位点（MERCs）内的蛋白质-蛋白质相互作用。©2025作者。Wiley期刊有限责任公司发表的当前协议。基本协议：骨骼肌组织和成肌细胞MERC蛋白的近端结扎试验。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Current protocols

CiteScore

4.00

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0.00%

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