Difference between revisions of "Permeabilized muscle fibers"
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The following references describe in detail the basis for working with muscle fibers in the O2k. | The following references describe in detail the basis for working with muscle fibers in the O2k. | ||
* [[Pesta | * [[Pesta 2012 Methods Mol Biol|Pesta D, Gnaiger E (2012) High-resolution respirometry. OXPHOS protocols for human cells and permeabilized fibres from small biopisies of human muscle. Methods Mol Biol 810: 25-58.]] | ||
* MiPNet14.14 PermeabilizedFibrePreparation| | * MiPNet14.14 PermeabilizedFibrePreparation|Preparation of permeabilized muscle fibres for diagnosis of mitochondrial respiratory function. Mitochondr Physiol Network 14.14.]] | ||
* [[MiPNet11.05 Mitos-PermeabilizedCells| | * [[MiPNet11.05 Mitos-PermeabilizedCells|Isolated mitochondria or permeabilized tissues and cells. Mitochondr. Physiol. Network 11.05.]] | ||
*[[MiPNet14.13 Medium-MiR06| | *[[MiPNet14.13 Medium-MiR06|Mitochondrial respiration medium - MiR06. Mitochondr Physiol Network 14.13.]] | ||
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Especially in muscle fibers an ADP concentration up to 5 mM needs to be applied to be in the saturating range, an ADP titration might be required to evaluate, whether the applied concentration is saturating. | Especially in muscle fibers an ADP concentration up to 5 mM needs to be applied to be in the saturating range, an ADP titration might be required to evaluate, whether the applied concentration is saturating. | ||
[[Pesta | [[Pesta 2012 Methods Mol Biol]]:At a high apparent Km for ADP of 0.5 mM, flux at 2.5 and 5 mM ADP is ADP-limited by 13% and 7% (assuming L/P=0.2). 2.5 mM ADP is saturating in many cases, yet a further increase of ADP concentration provides a test for saturating [ADP]. This is particularly important for evaluation of OXPHOS versus ETS capacity (P versus E; Fig. 5). | ||
== High oxygen concentration == | == High oxygen concentration == | ||
'''''Why is it necessary to measure muscle fiber respiration | '''''Why is it necessary to measure muscle fiber respiration above air saturation?''''' | ||
Respiratory flux of muscle fibre bundles is limited by oxygen diffusion even above 50% air saturation (O2 concentration of 100 ΞΌM). This problem is carefully controlled and avoided by application of a high oxygen regime. | Respiratory flux of muscle fibre bundles is limited by oxygen diffusion even above 50% air saturation (O2 concentration of 100 ΞΌM). This problem is carefully controlled and avoided by application of a high oxygen regime. | ||
'''See:''' [[Pesta | '''See:''' [[Pesta 2012 Methods Mol Biol]], [[Gnaiger 2003 Adv Exp Med Biol]]. | ||
Revision as of 20:28, 3 February 2012
References
The following references describe in detail the basis for working with muscle fibers in the O2k.
- Pesta D, Gnaiger E (2012) High-resolution respirometry. OXPHOS protocols for human cells and permeabilized fibres from small biopisies of human muscle. Methods Mol Biol 810: 25-58.
- MiPNet14.14 PermeabilizedFibrePreparation|Preparation of permeabilized muscle fibres for diagnosis of mitochondrial respiratory function. Mitochondr Physiol Network 14.14.]]
- Isolated mitochondria or permeabilized tissues and cells. Mitochondr. Physiol. Network 11.05.
- Mitochondrial respiration medium - MiR06. Mitochondr Physiol Network 14.13.
Chemicals and Media
What is the optimum ADP concentration to stimulate maximum muscle fibre respiration?
Especially in muscle fibers an ADP concentration up to 5 mM needs to be applied to be in the saturating range, an ADP titration might be required to evaluate, whether the applied concentration is saturating.
Pesta 2012 Methods Mol Biol:At a high apparent Km for ADP of 0.5 mM, flux at 2.5 and 5 mM ADP is ADP-limited by 13% and 7% (assuming L/P=0.2). 2.5 mM ADP is saturating in many cases, yet a further increase of ADP concentration provides a test for saturating [ADP]. This is particularly important for evaluation of OXPHOS versus ETS capacity (P versus E; Fig. 5).
High oxygen concentration
Why is it necessary to measure muscle fiber respiration above air saturation?
Respiratory flux of muscle fibre bundles is limited by oxygen diffusion even above 50% air saturation (O2 concentration of 100 ΞΌM). This problem is carefully controlled and avoided by application of a high oxygen regime.
See: Pesta 2012 Methods Mol Biol, Gnaiger 2003 Adv Exp Med Biol.
