SUIT-001 O2 ce-pce D004: Difference between revisions
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::: '''[[SUIT protocol pattern]]:''' orthogonal 1PM;2D;2c;3U;4G;5S;7Rot;8Gp- | ::: '''[[SUIT protocol pattern]]:''' orthogonal 1PM;2D;2c;3U;4G;5S;7Rot;8Gp- | ||
The SUIT-001 O2 ce-pce D004 protocol is the reference protocol 1 (RP1) specifically for PBMCs and PLTs. The main difference between the RP1 for PBMCs and PLTs and RP1 for other mitochondrial preparations [[SUIT-001| see SUIT-001]] is the absence of Oct in the case of blood cells. Β | The SUIT-001 O2 ce-pce D004 protocol is the reference protocol 1 (RP1) specifically for PBMCs and PLTs. The main difference between the RP1 for PBMCs and PLTs and RP1 for other [[Mitochondrial preparations| mitochondrial preparations]] ([[SUIT-001| see SUIT-001]]) is the absence of Oct in the case of blood cells. Β | ||
The SUIT-001 O2 ce-pce D004 protocol in combination with [[SUIT-002 O2 ce-pce D007a]] is specially designed to provide a common reference for comparison of respiratory control of [[PBMC|PBMC]] and [[Platelet|platelets]] cells (non-permeabilized cells must be added to the chambers and then permeabilized as a part of the protocol) in a wide variety of species. SUIT-001 O2 ce-pce D004 gives information of [[ROUTINE]] respiration (ce: non-permeabilized cells) and after the plasma membrane permeabilization (pce: permeabilized cells) of the linear coupling control ([[LEAK-respiration|''L'']]-[[Oxidative phosphorylation| ''P'']]-[[ET-capacity| ''E'']]) with NADH linked-substrates ([[PM pathway control state|PM]]). Moreover, the pathway control in ET state ([[NADH_Electron_transfer-pathway_state|N]], [[NS-pathway control state| NS]], [[Succinate pathway control state| S]] and [[SGp-pathway control state| SGp]] pathways) can be evaluated by using this SUIT protocol. SUIT-001 O2 ce-pce D004 can be extended with the CIV assay module. | The SUIT-001 O2 ce-pce D004 protocol in combination with [[SUIT-002 O2 ce-pce D007a]] is specially designed to provide a common reference for comparison of respiratory control of [[PBMC|PBMC]] and [[Platelet|platelets]] cells (non-permeabilized cells must be added to the chambers and then permeabilized as a part of the protocol) in a wide variety of species. SUIT-001 O2 ce-pce D004 gives information of [[ROUTINE]] respiration (ce: non-permeabilized cells) and after the plasma membrane permeabilization (pce: permeabilized cells) of the linear coupling control ([[LEAK-respiration|''L'']]-[[Oxidative phosphorylation| ''P'']]-[[ET-capacity| ''E'']]) with NADH linked-substrates ([[PM pathway control state|PM]]). Moreover, the pathway control in ET state ([[NADH_Electron_transfer-pathway_state|N]], [[NS-pathway control state| NS]], [[Succinate pathway control state| S]] and [[SGp-pathway control state| SGp]] pathways) can be evaluated by using this SUIT protocol. SUIT-001 O2 ce-pce D004 can be extended with the CIV assay module. | ||
Revision as of 16:41, 27 March 2019
Description
Abbreviation: NSGp(PGM)
Reference: A Reference protocol RP1 specific for PBMC and PTL cells; ce: non-permeabilized cells, pce: permeabilized cells - SUIT-001
SUIT number: D004_ce1;1Dig;1PM;2D;2c;3U;4G;5S;7Rot;8Gp;9Ama;10AsTm;11Azd
O2k-Application: O2
- MitoPedia: SUIT - SUIT reference protocol RP1 for PBMC and PLT cells that are permeabilized in the chamber
- SUIT-category: NSGp(PGM)
- SUIT protocol pattern: orthogonal 1PM;2D;2c;3U;4G;5S;7Rot;8Gp-
The SUIT-001 O2 ce-pce D004 protocol is the reference protocol 1 (RP1) specifically for PBMCs and PLTs. The main difference between the RP1 for PBMCs and PLTs and RP1 for other mitochondrial preparations ( see SUIT-001) is the absence of Oct in the case of blood cells. The SUIT-001 O2 ce-pce D004 protocol in combination with SUIT-002 O2 ce-pce D007a is specially designed to provide a common reference for comparison of respiratory control of PBMC and platelets cells (non-permeabilized cells must be added to the chambers and then permeabilized as a part of the protocol) in a wide variety of species. SUIT-001 O2 ce-pce D004 gives information of ROUTINE respiration (ce: non-permeabilized cells) and after the plasma membrane permeabilization (pce: permeabilized cells) of the linear coupling control (L- P- E) with NADH linked-substrates (PM). Moreover, the pathway control in ET state (N, NS, S and SGp pathways) can be evaluated by using this SUIT protocol. SUIT-001 O2 ce-pce D004 can be extended with the CIV assay module.
Communicated by Doerrier C and Gnaiger E (last update 2019-03-27)
Steps and respiratory states
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M) |
---|---|---|---|---|
ce1 | ROUTINE | ce1
| ||
1Dig | REN | ce1;1Dig
|
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M) |
---|---|---|---|---|
1PM | PML(n) | N | CI | PML or PM_L: Pyruvate & Malate, N-LEAK respiration, NL
|
2D | PMP | N | CI | 1PM;2D
|
2c | PMcP | N | CI | 1PM;2D;2c
|
3U | PME | N | CI | 1PM;2D;3U
|
4G | PGME | N | CI | 1PM;2D;3U;4G
|
5S | PGMSE | NS | CI&II | 1PM;2D;3U;4G;5S
|
(6Oct) | 1PM;2D;3U;4G;5S;(6Oct)
| |||
7Rot | SE | S | CII | 1PM;2D;3U;4G;5S;(6Oct);7Rot
|
8Gp | SGpE | SGp | CII&GpDH | 1PM;2D;3U;4G;5S;(6Oct);7Rot;8Gp
|
9Ama | ROX |
|
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
- Β» Electron transfer pathway
- Β» Fatty acid oxidation pathway control state, F
- Β» NADH electron transfer-pathway state, N
- Β» Succinate pathway control state, S
- Β» NS-pathway control state, NS
- Β» Glycerophosphate pathway control state, Gp
- Β» Complex IV single step, CIV
- Β» Anaplerotic pathway control state
- Pathway control
- Main fuel substrates
- Β» Glutamate, G
- Β» Glycerophosphate, Gp
- Β» Malate, M
- Β» Octanoylcarnitine, Oct
- Β» Pyruvate, P
- Β» 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 statistically used as repeated measurements.
- + In blood cells, SUIT-001 and SUIT-002 harmonize in: ROUTINE (ce1), SGpE (8Gp in SUIT-001 and 10Rot in SUIT-002) and Complex IV.
- + Harmonization with many SUIT protocols (up to step 5S).
- + This protocol can be extended with the Complex IV module.
- + The addition of non-permeabilized cells to the chambers provides additional information (ROUTINE respiration) which can not be obtained in SUIT-001 O2 mt D001 where the cells added to the chambers are already permeabilized.
- - The substrate combination for maximum ET capacity (FNSGP) is not obtained in SUIT-001 in favour of measuring SE.
- - Oct is not added in ET state.
- - Lengthy duration of the experiment.
Compare SUIT protocols
- SUIT-001 O2 ce-pce D004 (SUIT-001 for permeabilized PBMCs and PLTs) differs from SUIT-001 O2 ce-pce D003 (general SUIT-001 for permeabilized cells) in the absence and presence of octanoylcarnitine, respectively.
- SUIT-002 O2 ce-pce D07a (RP2): Harmonized SUIT protocol for PBMC and PLT cells (non-permeabilized cells must be added to the chambers and then permeabilized as a part of the protocol).
- SUIT-001 O2 mt D001: SUIT-001 for permeabilized cells (cells added to the chamber have been permeabilized before their addition). This SUIT protocol provides no information on ROUTINE respiration of non-permeabilized cells (ce).
References
Year | Reference | Organism | Tissue;cell | |
---|---|---|---|---|
MiPNet21.06 SUIT RP | 2018-06-25 | SUIT reference protocol for OXPHOS analysis by high-resolution respirometry. | ||
Doerrier 2018 Methods Mol Biol | 2018 | Doerrier C, Garcia-Souza LF, Krumschnabel G, Wohlfarter Y, MΓ©szΓ‘ros AT, Gnaiger E (2018) High-Resolution FluoRespirometry and OXPHOS protocols for human cells, permeabilized fibers from small biopsies of muscle, and isolated mitochondria. Methods Mol Biol 1782:31-70. https://doi.org/10.1007/978-1-4939-7831-1_3 | Human Mouse Rat Saccharomyces cerevisiae | Heart Skeletal muscle Endothelial;epithelial;mesothelial cell Blood cells HEK Platelet |
MitoPedia concepts:
SUIT protocol,
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MitoPedia methods:
Respirometry