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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