Difference between revisions of "Talk:MiPNet17.08 IOC68 Schroecken AT"
From Bioblast
(Created page with " Please add you Feedback here.") |
|||
(22 intermediate revisions by 5 users not shown) | |||
Line 1: | Line 1: | ||
== IOC68 Workshop Experiments == | |||
=== 2012-04-14 Demo Experiment === | |||
* Oxygraph AB (in both chambers the same SUIT protocol was applied) | |||
* DatLab File 2012-04-14_AB-02 | |||
* Mouse heart homogenate prepared with PBI Shredder (position 1 for 10 seconds, position 2 for 5 seconds) and the homogenate was completely washed out of the Shredder tube with 5 ml MiR05 | |||
* 2.3 mg tissue per chamber = 1.15 mg / ml | |||
* Flux derivation = 20 | |||
* Oxygen regime 320 to 160 (reoxygenation with pure oxygen at open chamber position) | |||
* Respiration medium: MiR05 (solubility factor = 0.92) | |||
* '''SUIT-Protocol''': PMG +AmR+HRP +H<sub>2</sub>O<sub>2</sub> +D2 +D3 +S +U(FCCP) +Rot +H<sub>2</sub>O<sub>2</sub> +H<sub>2</sub>O<sub>2</sub> +Mna +Ama | |||
* The homogenate was added, the oxygen concentration was increased up to 320 nmol/ml and then the O2k-chamber was closed. | |||
=== Notes === | |||
* The following respiratory states were obtained: | |||
:: CI-linked ''L'' respiration with 5 mM pyruvate, 2 mM malate and 10 mM glutamate in the absence of ADP (no adenylates, N) | |||
:: ADP-stimulated state ''P'' after titration of 3 mM ADP. | |||
:: CI<small>&</small>II-linked respiration after titration of 10 mM succinate. | |||
:: Noncoupled state ''E'' at 0.5 µM FCCP. | |||
:: S(Rot) (CII-linked) state ''E'' with 0.5 µM rotenone. | |||
* Titration of 3 mM ADP stimulated the oxygen flux about 69 to 73% and inhibited peroxide production by 1.0 to 4.7%. | |||
* Addition of succinate stimulated respiration in both chambers by about 27%, whereas H<sub>2</sub>O<sub>2</sub> flux was reduced by about 2.6 to 9.2%. | |||
* Reoxygenation with pure oxygen increased the O<sub>2</sub> flux (14 - 18%) as well as the H<sub>2</sub>O<sub>2</sub> flux (19 - 26%). | |||
* A direct influence of the O<sub>2</sub> concentration on induction of peroxide production occurred after reoxygenation with pure oxygen. | |||
* Increasing FCCP above 0.5 µM reduced the oxygen flux (2.7 – 6.6%) and the peroxide production (3.1 to 5 fold; Fig. 9A and B). In the end of the experiment, rotenone as inhibitor of C<sub>I</sub> was added, which reduced the oxygen flux (2 fold decrease) as well as the H<sub>2</sub>O<sub>2</sub> flux (25 - 29%) in both chambers. | |||
* In both chambers, the highest H<sub>2</sub>O<sub>2</sub> / O<sub>2</sub> flux ratio (''L'' with PMG) was 0.0034 (0.34%) and the addition of ADP diminished the H<sub>2</sub>O<sub>2</sub> / O<sub>2</sub> flux ratio to 0.0010 (0.10%). | |||
* Titration of succinate further diminished the ratio to 0.0007 (0.07%) and 0.5 µM FCCP resulted in a H<sub>2</sub>O<sub>2</sub> / O<sub>2</sub> flux ratio of 0.0002 (0.02%). | |||
* Reoxygenation caused an increase to 0.0007 (0.07%) and rotenone further increased the ratio to 0.0013 (0.13%). | |||
* Oxygen dependence of homogenate became visible as a decline of the oxygen flux and the H<sub>2</sub>O<sub>2</sub> flux together with the decline of the oxygen concentration occurred indicating a dependence of oxygen with homogenate that is, however, not as strong as observed with permeabilized fibres. | |||
* Generalized analysis of the experiments is difficult because of the effects of O<sub>2</sub> dependence. | |||
[[Image:BB-Bioblast.jpg|left|40px|link=Bioblast:About|Bioblast wiki]] | |||
== Popular Bioblast page == | |||
[[MiPNet17.08 IOC68 Schroecken AT]] has been accessed more than | |||
::::* 30,000 times (2019-07-22) | |||
::::* 25,000 times (2018-10-18) | |||
::::* 20,000 times (2016-01-26) | |||
::::* 15,000 times (2015-07-29) | |||
::::* 10,000 times (2014-07-18) | |||
::::* 5,000 times (2014-06-12) |
Latest revision as of 10:27, 22 July 2019
IOC68 Workshop Experiments
2012-04-14 Demo Experiment
- Oxygraph AB (in both chambers the same SUIT protocol was applied)
- DatLab File 2012-04-14_AB-02
- Mouse heart homogenate prepared with PBI Shredder (position 1 for 10 seconds, position 2 for 5 seconds) and the homogenate was completely washed out of the Shredder tube with 5 ml MiR05
- 2.3 mg tissue per chamber = 1.15 mg / ml
- Flux derivation = 20
- Oxygen regime 320 to 160 (reoxygenation with pure oxygen at open chamber position)
- Respiration medium: MiR05 (solubility factor = 0.92)
- SUIT-Protocol: PMG +AmR+HRP +H2O2 +D2 +D3 +S +U(FCCP) +Rot +H2O2 +H2O2 +Mna +Ama
- The homogenate was added, the oxygen concentration was increased up to 320 nmol/ml and then the O2k-chamber was closed.
Notes
- The following respiratory states were obtained:
- CI-linked L respiration with 5 mM pyruvate, 2 mM malate and 10 mM glutamate in the absence of ADP (no adenylates, N)
- ADP-stimulated state P after titration of 3 mM ADP.
- CI&II-linked respiration after titration of 10 mM succinate.
- Noncoupled state E at 0.5 µM FCCP.
- S(Rot) (CII-linked) state E with 0.5 µM rotenone.
- Titration of 3 mM ADP stimulated the oxygen flux about 69 to 73% and inhibited peroxide production by 1.0 to 4.7%.
- Addition of succinate stimulated respiration in both chambers by about 27%, whereas H2O2 flux was reduced by about 2.6 to 9.2%.
- Reoxygenation with pure oxygen increased the O2 flux (14 - 18%) as well as the H2O2 flux (19 - 26%).
- A direct influence of the O2 concentration on induction of peroxide production occurred after reoxygenation with pure oxygen.
- Increasing FCCP above 0.5 µM reduced the oxygen flux (2.7 – 6.6%) and the peroxide production (3.1 to 5 fold; Fig. 9A and B). In the end of the experiment, rotenone as inhibitor of CI was added, which reduced the oxygen flux (2 fold decrease) as well as the H2O2 flux (25 - 29%) in both chambers.
- In both chambers, the highest H2O2 / O2 flux ratio (L with PMG) was 0.0034 (0.34%) and the addition of ADP diminished the H2O2 / O2 flux ratio to 0.0010 (0.10%).
- Titration of succinate further diminished the ratio to 0.0007 (0.07%) and 0.5 µM FCCP resulted in a H2O2 / O2 flux ratio of 0.0002 (0.02%).
- Reoxygenation caused an increase to 0.0007 (0.07%) and rotenone further increased the ratio to 0.0013 (0.13%).
- Oxygen dependence of homogenate became visible as a decline of the oxygen flux and the H2O2 flux together with the decline of the oxygen concentration occurred indicating a dependence of oxygen with homogenate that is, however, not as strong as observed with permeabilized fibres.
- Generalized analysis of the experiments is difficult because of the effects of O2 dependence.
Popular Bioblast page
MiPNet17.08 IOC68 Schroecken AT has been accessed more than
- 30,000 times (2019-07-22)
- 25,000 times (2018-10-18)
- 20,000 times (2016-01-26)
- 15,000 times (2015-07-29)
- 10,000 times (2014-07-18)
- 5,000 times (2014-06-12)