In vivo measurements of respiration control by cytochrome c oxidase and in situ analysis of oxidative phosphorylation
Methods in cell biologyPublished 1 January 2001
Gaetano Villani, Giuseppe Attardi
Citations45
SJR quartileQ4
SJR score0.30
SNIP0.22
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TL;DR
This chapter proposes an approach determining the control that cytochrome c oxidase (COX) exerts on the rate of endogenous respiration in intact cell and a method employed for analysis of oxidative phosphorylation (OXPHOS) in permeabilized cells.
Abstract
This chapter proposes an approach determining the control that cytochrome c oxidase (COX) exerts on the rate of endogenous respiration in intact cell and a method employed for analysis of oxidative phosphorylation (OXPHOS) in permeabilized cells. The experimental procedures described are based on polarographic measurements of oxygen consumption by means of Clark-type oxygen electrodes. The equipment and its utilization are described in detail, and new generations of oxygen electrodes and oxygraphic chambers and software for the automatic analysis of respiratory kinetics are presented. With any chosen reaction chamber, it is important to measure the exact mixing volume. This corresponds to the minimal volume required to fill the chamber up to the bottom of the channel port of the stopper. This value is important for subsequent calculations. The volume introduced into the chamber has to exceed slightly the mixing volume in order to avoid formation of air bubbles from the vortex created by stirring.
Keywords
MedicineBiochemistry, Genetics and Molecular Biology
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