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Simultaneous Measurement of Mitochondrial Calcium and Mitochondrial Membrane Potential in Live Cells by Fluorescent Microscopy

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Fluorophore-Based Mitochondrial Ca²⁺ Uptake Assay.

Charles B Phillips^1,2

¹Department of Biochemistry, Brandeis University, Waltham, USA.

|August 16, 2021

Summary

This study presents a simple protocol to investigate mitochondrial calcium uptake using a calcium-sensitive dye. The method allows for unambiguous monitoring of extra-mitochondrial calcium levels in activated mammalian mitochondria.

Keywords:

Mitochondrial Calcium Flux HEK MCU Uniporter

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Area of Science:

Biochemistry
Cell Biology
Physiology

Background:

Mitochondrial calcium uptake is vital for cellular energy production, signaling, and programmed cell death.
Existing methods for studying mitochondrial calcium can be complex or yield ambiguous results.
A straightforward and reliable method is needed to investigate mitochondrial calcium dynamics.

Purpose of the Study:

To develop and present a simple, unambiguous protocol for investigating mitochondrial calcium uptake.
To enable researchers to easily monitor extra-mitochondrial calcium levels in activated mitochondria.

Main Methods:

The protocol utilizes a calcium-sensitive, membrane-impermeable fluorophore.
Mammalian cells with activated mitochondria are permeabilized for the experiment.
Extra-mitochondrial calcium levels are monitored using fluorescence measurements.

Main Results:

The developed protocol provides a straightforward method for studying mitochondrial calcium uptake.
The use of a specific fluorophore allows for unambiguous monitoring of calcium levels.
The method is applicable to permeabilized mammalian cells with activated mitochondria.

Conclusions:

This protocol offers a simple and effective approach to study the physiological importance of mitochondrial calcium.
The method facilitates unambiguous investigation of calcium dynamics in cellular processes.
The technique is valuable for researchers in cell biology and biochemistry.