SUIT-003 O2 ce D062: Difference between revisions
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:::* This protocol is aย [[Carrier control titrations|carrier control]] to [[SUIT-003 O2 ce D061]]. | :::* This protocol is aย [[Carrier control titrations|carrier control]] to [[SUIT-003 O2 ce D061]]. | ||
:::+ This is a cell [[coupling-control protocol]] that provides information on [[ROUTINE]] and [[Electron transfer pathway]] in the presence of ([[MitoKit-CII/Succinate-nv]]. | :::+ This is a cell [[coupling-control protocol]] that provides information on [[ROUTINE]] and [[Electron transfer pathway]] in the presence of ([[MitoKit-CII/Succinate-nv]]. | ||
:::- This protocol is provided without [[oligomycin]] addition. To also include [[LEAK]] in the protocol, an event for the addition of oligomycin and a related mark can be added by editing the DLP file, creating a [[Export DL-Protocol User (*.DLPU)|DLPU]]. However, the addition of [[oligomycin]] has to be carefully tested to avoid inhibitory effects in the evaluation of the [[ET | :::- This protocol is provided without [[oligomycin]] addition. To also include [[LEAK]] in the protocol, an event for the addition of oligomycin and a related mark can be added by editing the DLP file, creating a [[Export DL-Protocol User (*.DLPU)|DLPU]]. However, the addition of [[oligomycin]] has to be carefully tested to avoid inhibitory effects in the evaluation of the [[ET capacity]]. | ||
:::- Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler. | :::- Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler. | ||
Revision as of 16:07, 8 June 2020
Description
Abbreviation: CCP-ce Snv - control
Reference: A: Carrier control for the Coupling-control protocol for living cells with plasma membrane-permeable succinate (MitoKit-CII/Succinate-nv) - SUIT-003 O2 ce D050
SUIT number: D062_ce1;ce1DMSO;(ce2Omy);ce3U;ce4Rot;ce5Ama
O2k-Application: O2
- MitoPedia: SUIT - Coupling-control protocol (CCP) with the carrier of the plasma membrane-permeable succinate (MitoKit-CII/Succinate-nv) in living cells (ce): DMSO.
- SUIT protocol pattern: ce1;ce1DMSO;(ce2Omy);ce3U;ce4Rot;ce5Ama
SUIT-003 O2 ce D062 protocol is the control of the SUIT-003 O2 ce D050 adding the carrier DMSO in ROUTINE respiration instead of the plasma membrane-permeable succinate (MitoKit-CII/Succinate-nv). Moreover,the SUIT-003 O2 ce D062 protocol allows to evaluate the coupling control of living cells. Respiratory capacities are tested in a sequence of coupling states: ROUTINE and Electron transfer pathway. Optionally, to study LEAK respiration, the phosphorylation system might be inhibited by oligomycin. The final concentration of oligomycin has to be carefully optimized for various cell types, to minimize the inhibitory effect on the electron transfer system which would lead to an underestimation of ET capacity.
Communicated by Doerrier C (last update 2019-06-26)
Steps and respiratory states
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M)
|
---|---|---|---|---|
ce1 | ROUTINE | ce1
| ||
ce1DMSO | ROUTINE | ce1
| ||
(ce2Omy) | L(Omy) | ce1;ce1Snv;(ce2Omy)
| ||
ce3U | E | ce1;ce1Snv;(ce2Omy);ce3U
| ||
ce4Rot | ROX | ce1;ce1Snv;(ce2Omy);ce3U;ce4Rot
| ||
ce5Ama | ROX | ce1;ce1Snv;(ce2Omy);ce3U;ce4Rot;ce5Ama
|
- Bioblast links: SUIT protocols - >>>>>>> - Click on [Expand] or [Collapse] - >>>>>>>
- Coupling control
- Pathway control
- ยป Electron transfer pathway
- ยป Fatty acid oxidation pathway control state, F
- ยป NADH electron transfer-pathway state, N
- ยป Succinate pathway control state, S
- ยป NS-pathway control state, NS
- ยป Glycerophosphate pathway control state, Gp
- ยป Complex IV single step, CIV
- ยป Anaplerotic pathway control state
- Pathway control
- Main fuel substrates
- ยป Glutamate, G
- ยป Glycerophosphate, Gp
- ยป Malate, M
- ยป Octanoylcarnitine, Oct
- ยป Pyruvate, P
- ยป Succinate, S
- Main fuel substrates
- Glossary
Strengths and limitations
- This protocol is a carrier control to SUIT-003 O2 ce D061.
- + This is a cell coupling-control protocol that provides information on ROUTINE and Electron transfer pathway in the presence of (MitoKit-CII/Succinate-nv.
- - This protocol is provided without oligomycin addition. To also include LEAK in the protocol, an event for the addition of oligomycin and a related mark can be added by editing the DLP file, creating a DLPU. However, the addition of oligomycin has to be carefully tested to avoid inhibitory effects in the evaluation of the ET capacity.
- - Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler.
Compare SUIT protocols
- SUIT-003 O2 ce D050 the plasma membrane-permeable succinate (MitoKit-CII/Succinate-nv) is added in ROUTINE.
References
MitoPedia concepts: SUIT protocol, SUIT B, Find
MitoPedia methods:
Respirometry