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Updated: May 25, 2026
Imaging and Quantifying Mitochondrial Morphology in C. elegans During Aging
Published on: January 17, 2025
Protocol for the in vivo quantification of mitochondria-associated ER membranes in Caenorhabditis elegans
Fivos Borbolis1, Konstantinos Palikaras2
1Deparment of Biology, University of Patras, Patras 26504, Greece.
Abstract:
Mitochondria-associated membranes (MAMs) are specialized contact sites between the endoplasmic reticulum and mitochondria, with multiple functional aspects. Here, we present a confocal microscopy-based protocol for quantifying the area of MAM-like domains in intestinal cells of the nematode Caenorhabditis elegans using organelle-specific fluorescent reporters. We describe steps for worm synchronization, microscope setup, sample preparation, and image acquisition. We then detail procedures for manual single-image analysis and scalable automated batch processing, enabling robust quantification of ER-mitochondria contacts across experimental conditions. For complete details on the use and execution of this protocol, please refer to Roussos et al.1.
Insights
This study introduces a confocal microscopy protocol to measure mitochondria-associated membranes (MAMs) in C. elegans intestinal cells. The method enables robust quantification of endoplasmic reticulum-mitochondria contacts for research.
Area of Science:
- Cell Biology
- Organelle Biology
- Microscopy Techniques
Background:
- Mitochondria-associated membranes (MAMs) are critical contact sites between the endoplasmic reticulum and mitochondria.
- MAMs play diverse roles in cellular functions, necessitating accurate quantification methods.
- Understanding ER-mitochondria contacts is key to deciphering cellular processes.
Purpose of the Study:
- To present a detailed confocal microscopy protocol for quantifying MAM-like domains.
- To enable robust measurement of endoplasmic reticulum-mitochondria contacts in C. elegans.
- To provide a method for scalable analysis of MAMs across experimental conditions.
Main Methods:
- Utilized confocal microscopy with organelle-specific fluorescent reporters in C. elegans intestinal cells.
- Developed a protocol covering worm synchronization, sample preparation, and image acquisition.
- Detailed manual and automated image analysis procedures for quantifying MAM domain area.
Main Results:
- Successfully established a reproducible protocol for quantifying MAM-like domains.
- Demonstrated the utility of the protocol for robust quantification of ER-mitochondria contacts.
- The method allows for scalable batch processing of images for high-throughput analysis.
Conclusions:
- The presented protocol offers a reliable method for assessing ER-mitochondria contact sites.
- This technique facilitates quantitative studies of MAMs in C. elegans.
- The protocol supports diverse research applications investigating mitochondrial and ER function.
