- high-resolution terminology - matching measurements at high-resolution
SUIT-006 NADH mt D084
Description
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Reference: A: protocol for simultaneous determination of O2 flux and NADH autofluorescence in mitochondrial preparations (isolated mitochondria, tissue homogenate and permeabilized cells)- SUIT-006
SUIT number: D084_1PGM;2D;3U;3anoxia;4Myx
O2k-Application: NADH
Communicated by Doerrier C (last update 2022-05-03)
Representative traces
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Steps and respiratory states
Step
|
State
|
Pathway
|
Q-junction
|
Comment - Events (E) and Marks (M)
|
mt
|
REN
|
|
|
mt
- Respiration in the REN state is due to the presence of residual endogenous substrates.
- In the absence of endogenous substrates, mt can be used for calibration of fully oxidized NAD.
|
1PGM
|
PGML(n)
|
N
|
CI
|
1PGM
|
2D
|
PGMP
|
N
|
CI
|
1PGM;2D
|
(3Omy)
|
PGML(Omy)
|
N
|
CI
|
1PGM;2D;(3Omy)
- Omy addition is skipped in SUIT-006 NADH mt D084
- NADH-linked substrates (type N-pathway to Q).
- Non-phosphorylating resting state (LEAK state); LEAK-respiration, L(Omy), after blocking the ATP synthase with oligomycin.
|
4U
|
PGME
|
N
|
CI
|
1PGM;2D;(3Omy);4U
|
5Anox
|
|
N
|
CI
|
1PGM;2D;(3Omy);4U;5Anox
- Anoxia, after the biological sample has consumed the O2 in the O2k-chamber, is used to detect the fully reduced NAD for calculation of the NAD redox states.
|
6Myx
|
|
N
|
CI
|
1PGM;2D;(3Omy);4U;5Anox;6Myx
- The addition of myxothiazol after anoxia is a crucial step to detect the fully reduced NAD for calculation of the NAD redox state. Myxothiazol can induce a further reduction of NAD under anoxia.
|
7Reox
|
ROX
|
|
|
1PGM;2D;(3Omy);4U;5Anox;6Myx;7Reox
- The reoxygenation by opening the chamber in the presence of Myx allows for Rox-correction of the O2 fluxes.
|
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- Coupling control
- » Coupling control state
- » ET capacity
- » OXPHOS capacity
- » LEAK respiration
- Pathway control
- » Electron transfer pathway
- » Fatty acid oxidation pathway control state, F
- » NADH electron transfer-pathway state, N
- » Succinate pathway control state, S
- » NS-pathway control state, NS
- » Glycerophosphate pathway control state, Gp
- » Complex IV single step, CIV
- » Anaplerotic pathway control state
- Main fuel substrates
- » Glutamate, G
- » Glycerophosphate, Gp
- » Malate, M
- » Octanoylcarnitine, Oct
- » Pyruvate, P
- » Succinate, S
- Glossary
- » List of SUIT states
- » SUIT concept
Strengths and limitations
Compare SUIT protocols
Chemicals and syringes
- Suggested stock concentrations are shown in the specific DL-Protocol.
References