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Measurement of mitochondrial membrane potential in cardiomyocytes

FJ Feifan Jia
YC Yuanyuan Chen
GX Gaojie Xin
LL Lingmei Li
ZL Zixin Liu
SX Sujuan Xu
JG Jiaming Gao
HM Hongxu Meng
YS Yue Shi
YM Yanlei Ma
LL Lei Li
JF Jianhua Fu
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A portion of the myocardium was excised and weighed, rinsed in saline and then washed twice in 1 ml of Lysis Buffer. After 20 grinding sessions in an ice bath at 0 °C, the tissue was homogenised and divided into centrifuge tubes and centrifuged at 1000 g for 5 min at 4 °C. The supernatant was taken out and placed to a new centrifuge tube and centrifuge again for 5 min. Next, transfer to a new tube and centrifuge at 12,000 g for 10 min at 4 °C. Let the mitochondria settle at the bottom of the tube after removing the supernatant. Add 0.5 mL Wash Buffer to resuspend the precipitate, centrifuge at 1000 g for 5 min at 4 °C and then centrifuge again at 12000 g for 10 min at 4 °C. Discard the supernatant and the precipitate will be high purity mitochondria. The precipitate can be resuspended with 50–100 μL of Store Buffer or reaction buffer and used immediately to detect the mitochondrial membrane potential using the JC-1 fluorescent probe method.

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