Vienne 2017 Anal Biochem
|Vienne JC, Cimetta C, Dubois M, Duburcq T, Favory R, Dessein AF, Fontaine M, Joncquel-Chevalier Curt M, Cuisset JM, Douillard C, Mention-Mulliez K, Dobbelaere D, Vamecq J (2017) A fast method for high resolution oxymetry study of skeletal muscle mitochondrial respiratory chain complexes. Anal Biochem 528:57-62.|
Abstract: High resolution oxymetry study (HROS) of skeletal muscle usually requires 90-120 min preparative phase (dissection, permeabilization and washing). This work reports on the suitability of a rapid muscle preparation which by-passes this long preparation. For a few seconds only, muscle biopsy from pigs is submitted to gentle homogenization at 8000 rotations per minute using an ultra-dispersor apparatus. Subsequent HROS is performed using FCCP instead of ADP, compounds crossing and not plasma membrane, respectively. This simplified procedure compares favorably with classical (permeabilized fibers) HROS in terms of respiratory chain complex activities. Mitochondria from cells undergoing ultradispersion were functionally preserved as attested by relative inefficacy of added cytochrome C (not crossing intact mitochondrial outer membrane) to stimulate mitochondrial respiration. Responsiveness of respiration to ADP (in the absence of FCCP) suggested that these intact mitochondria were outside cells disrupted by ultradispersion or within cells permeated by this procedure.
Copyright © 2017 Elsevier Inc. All rights reserved. • Keywords: electron transfer-pathway (ET-pathway), FCCP, High resolution oxymetry, MIR05 buffer, Muscle needle biopsy, Respiratory chain complex, Saponin-permeabilized fibers, Ultradispersion • Bioblast editor: Kandolf G • O2k-Network Lab: FR Lille Boutry M, FR Lille Vienne JC
Labels: MiParea: Respiration, Instruments;methods
Organism: Pig Tissue;cell: Skeletal muscle Preparation: Permeabilized tissue, Homogenate
Coupling state: LEAK, OXPHOS, ET Pathway: N, S, CIV, ROX HRR: Oxygraph-2k