Mitochondrial Ca homeostasis is crucial for regulating vital functions such as respiration or apoptosis. Targeted aequorins are excellent probes to measure subcellular Ca. Ca concentration in mitochondria ([Ca]) is low at rest (about 10 M) and can increase to the micromolar or even approach the millimolar range, upon cell activation. Here we describe a new quantitative luminescent protocol to directly measure mitochondrial Ca uptake, optimized for high throughput. The sensitivity of the method allows detection of changes in either the capacity or the affinity of mitochondrial Ca transport.
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