From Bioblast
Description
Abbreviation: ceGlc
Reference: A Coupling control protocol and Crabtree effect test -SUIT-003
SUIT number: D038_ce1;ce2(Omy);ce3U;ce3Glc;ce3(U);ce4Ama
O2k-Application: O2
- MitoPedia: SUIT
- SUIT-category: ceGlc
- SUIT protocol pattern: complex cell coupling control protocol ce1;ce2(Omy);ce3U;ce3Glc;ce3(U);ce4Ama
SUIT-003 O2 ce D038 has been designed to study the coupling control state and respiration regulation by glucose (Crabtree effect) of intact cells on ET-capacity. To obtain more information about Crabtree effect, this protocol can be used in parallel with SUIT-003 O2 ce D037. SUIT-003 O2 ce D038 follows a modular approach and can be easily used in combination with the assessment of complex IV capacity and respirometric cell viability test.
Communicated by Garcia-Souza LF and Cardoso LH (last update 2019-03-28)
Steps and respiratory states
Step | State | Pathway | Q-junction | Comment - Events (E) and Marks (M) |
---|---|---|---|---|
ce1 | ROUTINE | ce1
| ||
(ce2Omy) | L(Omy) | ce1;(ce2Omy)
| ||
ce3U | E | ce1;(ce2Omy);ce3U
| ||
ce3Glc | E | ce1;(ce2Omy);ce3U;ce3Glc
| ||
ce3'U | E | ce1;(ce2Omy);ce3U;ce3Glc;ce3'U
| ||
ce4Ama | ROX | ce1;(ce2Omy);ce3U;ce3Glc;ce3'U;ce4Ama
|
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- 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
- The SUIT-003 O2 ce D038 is a cell coupling control protocol. The total cell count (Nce) is the sum of viable cells (Nvce) and dead cells (Ndce).
- This protocol can be extended with the cell viability test module (See CCV SUIT-003 O2 ce-pce D020).
- CIV activity can also be measured after the cell viability test module.
- + Reasonable duration of the experiment.
- + To obtain more information about Crabtree effect, this protocol can be used in parallel with SUIT-003 O2 ce D037. In SUIT-003 O2 ce D037, the addition of glucose during ROUTINE state can lead to increase in ATP production by glycolysis, which can indirectly decrease respiration. In SUIT-003 O2 ce D038, with addition of glucose in ET state, this effect is bypassed, and only the direct effect of glucose on the electron-transfer system is tested.
- - Cell culture media with glucose (e.g. DMEM: 25 Β΅M Glucose) could hinder the cell viability test.
- - Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler.
Compare SUIT protocols
- SUIT-003 O2 ce D037 can be used in parallel to obtain more information on Crabtree effect.
- SUIT-003 O2 ce-pce D013 for assessment of coupling control, Crabtree effect and cell viability test.
- SUIT-003 O2 ce-pce D018 for assessment of coupling control, Crabtree effect, malate effect and cell viability test.
References
MitoPedia concepts: SUIT protocol, SUIT A, Find
MitoPedia methods:
Respirometry