Difference between revisions of "SUIT-015 O2 pti D043"
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|info='''A''' pti: permeabilized tissue- '''[[SUIT-015]]''' | |info='''A''' pti: permeabilized tissue- '''[[SUIT-015]]''' | ||
|abbr=FNS(Oct,PGM) | |abbr=FNS(Oct,PGM) | ||
|SUIT number=D043_1OctM;2D;3G;4P;5S;6U;7Rot;8Ama | |SUIT number=D043_1OctM;2D;2c;3G;4P;5S;6U;7Rot;8Ama | ||
}} | }} | ||
::: '''[[Categories of SUIT protocols |SUIT-category]]:''' FNS(Oct,PGM) | ::: '''[[Categories of SUIT protocols |SUIT-category]]:''' FNS(Oct,PGM) |
Latest revision as of 11:41, 29 February 2024
Description
Abbreviation: FNS(Oct,PGM)
Reference: A pti: permeabilized tissue- SUIT-015
SUIT number: D043_1OctM;2D;2c;3G;4P;5S;6U;7Rot;8Ama
O2k-Application: O2
- SUIT-category: FNS(Oct,PGM)
- SUIT protocol pattern: diametral 1OctM;2D;3G;4P;5S;6U:7Rot-
SUIT-015 O2 pti D043 gives information on F-pathway in LEAK state and OXPHOS state avoiding FAO overestimation in the presence of anaplerotic pathways. In addition, the pathway control of FN and FNS in OXPHOS state and of FNS and S in ET state can be evaluated. SUIT-015 O2 pti D043 can be extended with the CIV assay module. Permeabilized muscle fibers are sensitive to oxygen supply due to limited diffusion of oxygen to the fiber bundle core. To counteract this limitation, hyperoxic conditions (400-250 µM O2) must be employed. To set the optimal oxygen concentration in the O2k-Chamber, see Setting the oxygen concentration.
Communicated by Gnaiger E (last update 2019-02-04)
Representative traces
Steps and respiratory states
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M) |
---|---|---|---|---|
1OctM | OctML(n) | F(N) | CETF | 1OctM
|
2D | OctMP | F(N) | CETF | 1OctM;2D
|
3G | OctGMP | FN | CETF&CI | 1OctM;2D;3G
|
4P | OctPGMP | FN | CETF&CI | 1OctM;2D;3G;4P
|
5S | OctPGMSP | FNS | CETF&CI&II | 1OctM;2D;3G;4P;5S
|
6U | OctPGMSE | FNS | CETF&CI&II | 1OctM;2D;3G;4P;5S;6U
|
7Rot | SE | S | CII | 1OctM;2D;3G;4P;5S;6U;7Rot
|
8Ama | ROX | 1OctM;2D;3G;4P;5S;6U;7Rot;8Ama
|
Step | Respiratory state | Pathway control | ET-Complex | Comment |
---|---|---|---|---|
## AsTm | AsTmE | CIV | CIV | |
## Azd | CHB |
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- Coupling control
- Pathway control
- Main fuel substrates
- » Glutamate, G
- » Glycerophosphate, Gp
- » Malate, M
- » Octanoylcarnitine, Oct
- » Pyruvate, P
- » Succinate, S
- Main fuel substrates
- Glossary
Strengths and limitations
- + The protocol provides information on FAO capacity in the absence of other, potentially interfering pathways, both in the LEAK state and in OXPHOS.
- + FNS OXPHOS capacity comprises the most important pathways in many cell types and thus provides a physiologically relevant estimate of maximum mitochondrial respiratory capacity.
- + FNS ET capacity is a good estimate of overall ET capacity in many cell types.
- + The presence of PMG and S establishes a fully operative TCA cycle activity.
- + Reasonable duration of the experiment.
- - F OXPHOS capacity may be underestimated. In human heart muscle addition of Oct to palmitoylcarnitine (Pal) + malate increased OXPHOS by 26% (Lemuieux et al 2011).
- - SRot(E) may be underestimated if S is not saturating.
- - CIV activity is not measured, to save experimental time.
Compare SUIT protocols
Chemicals and syringes
Step | Chemical(s) and link(s) | Comments |
---|---|---|
1OctM | Octanoylcarnitine (Oct) and Malate (M) | |
2D | ADP (D) | |
3G | Glutamate (G) | |
4P | Pyruvate (P) | |
5S | Succinate (S) | |
6U | Carbonyl cyanide m-chlorophenyl hydrazone, CCCP (U) | Can be substituted for other uncoupler |
7Rot | Rotenone (Rot) | |
8Ama | Antimycin A (Ama) |
- Suggested stock concentrations are shown in the specific DL-Protocol.
References
Year | Reference | Organism | Tissue;cell | |
---|---|---|---|---|
Schoepf 2016 FEBS J | 2016 | Schöpf B, Schäfer G, Weber A, Talasz H, Eder IE, Klocker H, Gnaiger E (2016) Oxidative phosphorylation and mitochondrial function differ between human prostate tissue and cultured cells. https://doi.org/10.1111/febs.13733 | Human | Endothelial;epithelial;mesothelial cell Genital Other cell lines Fibroblast |
MitoPedia concepts: SUIT protocol, SUIT A, Find
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Respirometry