Difference between revisions of "SUIT-004 O2 pfi D010"
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::::* [[Permeabilized muscle fibres |pfi]], high O<sub>2</sub>, no ROX: NS(PM)01_pfiO2,1PM,2D(c),3U,4S,5Rot | ::::* [[Permeabilized muscle fibres |pfi]], high O<sub>2</sub>, no ROX: NS(PM)01_pfiO2,1PM,2D(c),3U,4S,5Rot | ||
:::* | ::::* The '''[[SUIT-004]]''' protocol is linked to [[SUIT-001 O2 pfi D002]] for healthy human skeletal muscle mitochondria on the basis of the following rationale. | ||
::::# The first coupling control steps (1-4) are identical. | ::::# The first coupling control steps (1-4) are identical. |
Revision as of 16:35, 22 January 2019
Description
Abbreviation: NS(PM)
Reference: A: SUIT-004 short protocol linked to SUIT-001 O2 pfi D002 - SUIT RP1 (human skeletal muscle)
SUIT number: D010_1PM;2D;3U;4S;5Rot
O2k-Application: O2
- SUIT-category: NS(PM)
- SUIT protocol pattern: orthogonal 1PM;2D;2c;3U;4S;5Rot
- Linked to SUIT-001 O2 pfi D002 - SUIT RP1, specifically for human skeletal muscle mitochondria.
- Harmonized with SUIT-005 O2 pfi D011.
References
Steps and respiratory states
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M) |
---|---|---|---|---|
1PM | PML(n) | N | CI | 1PM
|
2D | PMP | N | CI | 1PM;2D
|
2c | PMcP | N | CI | 1PM;2D;2c
|
3U | PME | N | CI | 1PM;2D;3U
|
4S | PMSE | NS | CI&II | 1PM;2D;3U;4S
|
5Rot | SE | S | CII | 1PM;2D;3U;4S;5Rot
|
6Ama | ROX | 1PM;2D;3U;4S;5Rot;6Ama
|
Step | Respiratory state | Pathway control | ET-Complex | Comment |
---|---|---|---|---|
## AsTm | AsTmE | CIV | CIV | |
## Azd | CHB |
- Bioblast links: SUIT protocols - >>>>>>> - Click on [Expand] or [Collapse] - >>>>>>>
- 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
- SUIT-001 in combination with SUIT-002 provides a common reference for comparison of respiratory control in a large variety of species, tissues and cell types. Both SUIT protocols provide a mitochondrial mapping which allows:
- 1. to obtain reference values.
- 2. to evaluate mitochondrial physiological diversity, generating a mt-database on comparative mitochondrial physiology.
- 3. to screen specific defects.
- SUIT-001 and SUIT-002 are used in the MitoFit Proficiency test for inter-individual and inter-laboratory reproducibility quality control.
- A succinate concentration of >10 mM may be required for saturating SE capacity.
- + Linear coupling control (L-P-E) with N substrates (PM).
- + Pathway control in ET state (N, NS, FNS, S and SGp pathways).
- + Harmonization with SUIT-002 allows to perform both SUIT protocols in parallel. The cross-linked respiratory states can be statically used as repeat measurements.
- + In mtprep, SUIT-001 and SUIT-002 harmonize in: SGpE (8Gp in SUIT-001 and 10Rot in SUIT-002) and Complex IV.
- + Harmonization with many SUIT protocols (up to step 5S).
- + Oct is added in ET state to avoid uncoupler effect with fatty acids.
- + If NS- and FNS-pathways are similar (NSE ~ FNSE), the additivity index of N- and S-pathways in ET state can be evaluated.
- + This protocol can be extended with the Complex IV module.
- - The substrate combination for maximum ET capacity (FNSGP) is not obtained in SUIT-001 in favour of measuring SE.
Compare SUIT protocols
- The SUIT-004 protocol is linked to SUIT-001 O2 pfi D002 for healthy human skeletal muscle mitochondria on the basis of the following rationale.
- The first coupling control steps (1-4) are identical.
- RP1-5G is skipped, since PM and PGM yield nearly identical OXPHOS capacity. However, PM<PGM is a potential diagnostic for a defect in the PM-linked pathway upstream of CI.
- RP1-7Oct is skipped, since octanoylcarnitine exerts practically no stimulatory effect on OXPHOS capacity in state PMS or PGMS, but in conjunction with prolonged exposure may even yield a slight decline of respiration.
- RP1-9Gp is skipped, since glycerolphosphate exerts only a minor stimulatory effect on OXPHOS capacity in state SRot.
- Omission of the three steps described above shortens the SUIT protocol, thus reducing the risk of diminishing respiratory capacity over time, reducing the number of necessary reoxygenations, and providing more time for detailed evaluation of steady-states. The loss of diagnostic information may be minor relative to the benefits of simplification of the protocol, particularly in studies of exercise in healthy humans.
- RP1-10Ama may be skipped, since correction of the initial states by ROX measured in RP1-10 Ama may represent an over-correction, and inhibition takes a very long time in several cases (Pesta 2012 Methods Mol Biol).
- RP1-11Tm and RP1-12Azd are skipped, if a prolonged protocol is not practical due to limination of time.
MitoPedia concepts:
SUIT protocol,
SUIT A,
Find
MitoPedia methods:
Respirometry