Preparation and respirometric assessment of mitochondria isolated from skeletal muscle tissue obtained by percutaneous needle biopsy

Journal of Visualized Experiments : JoVE
Manish S BharadwajAnthony J A Molina

Abstract

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining samples of human Vastus lateralis, isolating mitochondria from minimal amounts of skeletal muscle tissue, and plate based respirometric profiling using an extracellular flux (XF) analyzer. Comparison of respirometric profiles obtained using 1.0, 2.5 and 5.0 μg of mitochondria indicate that 1.0 μg is sufficient to measure respiration and that 5.0 μg provides most consistent results based on comparison of standard errors. Western blot analysis of isolated mitochondria for mitochondrial marker COX IV and non-mitochondrial tissue marker GAPDH indicate that there is limited non-mitochondrial contamination using this protocol. The ability to study mitochondrial respirometry in as little as 20 mg of muscle tissue allows users to utilize individual biopsies for multiple study endpoints in clinical research projects.

Citations

Sep 3, 2017·European Journal of Applied Physiology·Laura C O'BrienAshraf S Gorgey
Aug 26, 2019·Reproductive Biology and Endocrinology : RB&E·Hannah M AtkinsDavid L Caudell
May 18, 2019·Archives of Pathology & Laboratory Medicine·Kenneth P H Pritzker, Heikki J Nieminen
Nov 4, 2017·Oxidative Medicine and Cellular Longevity·Daniel J TyrrellAnthony J A Molina
Sep 13, 2019·Nutrients·Lipika SalayeDonald A McClain
May 21, 2021·The Journal of General Physiology·Yoel H SitbonDanuta Szczesna-Cordary

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Methods Mentioned

BETA
biopsies
biopsy
Assay
dissecting
Protein Assay

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