Difference between revisions of "MiPNet17.17 Amplex-Mouse-brain"
Line 1: | Line 1: | ||
{{Publication | {{Publication | ||
|title=[[Image:O2k-Protocols.jpg|right|80px|link=http:// | |title=[[Image:O2k-Protocols.jpg|right|80px|link=http://wiki.oroboros.at/index.php/O2k-Protocols|O2k-Protocols]] O2k-Fluorometry: HRR and H<sub>2</sub>O<sub>2</sub> production in mouse brain mitochondria. | ||
|info=[[Media:MiPNet17.17_Amplex-Mouse-brain.pdf|'''Open Access-MiPNet17.17.pdf''']] ;[http://www.bioblast.at/index.php/File:MiPNet17.17_Amplex-Mouse-brain.pdf Versions] | |info=[[Media:MiPNet17.17_Amplex-Mouse-brain.pdf|'''Open Access-MiPNet17.17.pdf''']] ;[http://www.bioblast.at/index.php/File:MiPNet17.17_Amplex-Mouse-brain.pdf Versions] | ||
|authors=OROBOROS | |authors=OROBOROS | ||
|year=2013-10 | |year=2013-10 | ||
|journal=Mitochondr Physiol Network | |journal=Mitochondr Physiol Network | ||
|abstract='''O2k- | |abstract='''Fasching M, Sumbalova Z, Gnaiger E (2013) O2k-Fluorometry: HRR and H<sub>2</sub>O<sub>2</sub> production in mouse brain mitochondria. Mitochondr Physiol Network 17.17.''' | ||
High-resolution respirometry [1] is extended by MultiSensor techniques allowing the simultaneous measurement of oxygen consumption and one additional parameter (mitochondrial membrane potential, pH, Ca<sup>2+</sup>, or NO concentration [2-5]). Fluorometric methods are available for a wide range of analytical parameters of major interest in mitochochondrial physiology: H<sub>2</sub>O<sub>2</sub> production, mitochondrial membrane potential, intracellular pH, Ca<sup>2+</sup>, Mg<sup>2+</sup>, and NADH levels. | [[High-resolution respirometry]] [1] is extended by MultiSensor techniques allowing the simultaneous measurement of oxygen consumption and one additional parameter (mitochondrial membrane potential, pH, Ca<sup>2+</sup>, or NO concentration [2-5]). Fluorometric methods are available for a wide range of analytical parameters of major interest in mitochochondrial physiology: H<sub>2</sub>O<sub>2</sub> production, mitochondrial membrane potential, intracellular pH, Ca<sup>2+</sup>, Mg<sup>2+</sup>, and NADH levels. | ||
Advantages of the simultaneous measurement of additional parameters in a single respirometric chamber: | Advantages of the simultaneous measurement of additional parameters in a single respirometric chamber: | ||
# Quality control of preparations by respirometric performance. | # Quality control of preparations by respirometric performance. | ||
Line 15: | Line 15: | ||
# Oxygen dependence of parameters can be studied. | # Oxygen dependence of parameters can be studied. | ||
Here we introduce the O2k-Fluorescence LED2-Module, describe one selected stage in its development, and show its application for the simultaneous measurement of respiration and H<sub>2</sub>O<sub>2</sub> production of mouse brain mitochondria. | Here we introduce the O2k-Fluorescence LED2-Module, describe one selected stage in its development, and show its application for the simultaneous measurement of respiration and H<sub>2</sub>O<sub>2</sub> production of mouse brain mitochondria. | ||
:>> Product: [http://www.oroboros.at/?oxygraph OROBOROS Oxygraph-2k], [[O2k-Catalogue_OROBOROS| O2k-Catalogue]] | :>> Product: [http://www.oroboros.at/?oxygraph OROBOROS Oxygraph-2k], [[O2k-Catalogue_OROBOROS| O2k-Catalogue]] | ||
|mipnetlab=AT_Innsbruck_OROBOROS, AT Innsbruck MitoCom | |mipnetlab=AT_Innsbruck_OROBOROS, AT Innsbruck MitoCom}} | ||
}} | |||
{{Labeling | {{Labeling | ||
|area=Respiration, Instruments;methods | |area=Respiration, Instruments;methods | ||
Line 29: | Line 26: | ||
|substratestates=CI, CII, CIII, ROX | |substratestates=CI, CII, CIII, ROX | ||
|instruments=Oxygraph-2k, Fluorometry, Protocol | |instruments=Oxygraph-2k, Fluorometry, Protocol | ||
|additional=O2k-Demo | |additional=O2k-Demo}} | ||
}} | |||
== Further Information == | == Further Information == | ||
Revision as of 15:51, 17 May 2014
O2k-Fluorometry: HRR and H2O2 production in mouse brain mitochondria. |
ยป Open Access-MiPNet17.17.pdf ;Versions
OROBOROS (2013-10) Mitochondr Physiol Network
Abstract: Fasching M, Sumbalova Z, Gnaiger E (2013) O2k-Fluorometry: HRR and H2O2 production in mouse brain mitochondria. Mitochondr Physiol Network 17.17.
High-resolution respirometry [1] is extended by MultiSensor techniques allowing the simultaneous measurement of oxygen consumption and one additional parameter (mitochondrial membrane potential, pH, Ca2+, or NO concentration [2-5]). Fluorometric methods are available for a wide range of analytical parameters of major interest in mitochochondrial physiology: H2O2 production, mitochondrial membrane potential, intracellular pH, Ca2+, Mg2+, and NADH levels. Advantages of the simultaneous measurement of additional parameters in a single respirometric chamber:
- Quality control of preparations by respirometric performance.
- Objective exclusion criteria for protocol related problems, e.g. overtitration of uncouplers, side effects of TPP+ or fluorophores.
- Elimination of artifacts of normalization for a mitochondrial marker when combining two methods in a single chamber.
- Additional information is acquired for a limited amount of sample.
- Oxygen dependence of parameters can be studied.
Here we introduce the O2k-Fluorescence LED2-Module, describe one selected stage in its development, and show its application for the simultaneous measurement of respiration and H2O2 production of mouse brain mitochondria.
- >> Product: OROBOROS Oxygraph-2k, O2k-Catalogue
โข O2k-Network Lab: AT_Innsbruck_OROBOROS, AT Innsbruck MitoCom
Labels: MiParea: Respiration, Instruments;methods
Organism: Mouse
Tissue;cell: Nervous system
Preparation: Isolated Mitochondria"Isolated Mitochondria" is not in the list (Intact organism, Intact organ, Permeabilized cells, Permeabilized tissue, Homogenate, Isolated mitochondria, SMP, Chloroplasts, Enzyme, Oxidase;biochemical oxidation, ...) of allowed values for the "Preparation" property.
Regulation: Uncoupler Coupling state: LEAK, OXPHOS, ETS"ETS" is not in the list (LEAK, ROUTINE, OXPHOS, ET) of allowed values for the "Coupling states" property.
HRR: Oxygraph-2k, Fluorometry"Fluorometry" is not in the list (Oxygraph-2k, TIP2k, O2k-Fluorometer, pH, NO, TPP, Ca, O2k-Spectrophotometer, O2k-Manual, O2k-Protocol, ...) of allowed values for the "Instrument and method" property., Protocol"Protocol" is not in the list (Oxygraph-2k, TIP2k, O2k-Fluorometer, pH, NO, TPP, Ca, O2k-Spectrophotometer, O2k-Manual, O2k-Protocol, ...) of allowed values for the "Instrument and method" property.
O2k-Demo
Further Information
References
- O2k-Publications: O2k-Fluorometry
- Pesta D, Gnaiger E (2012) High-resolution respirometry. OXPHOS protocols for human cells and permeabilized fibres from small biopisies of human muscle. Methods Mol Biol 810: 25-58.
- Aguirre E, Rodrรญguez-Juรกrez F, Bellelli A, Gnaiger E, Cadenas S (2010) Kinetic model of the inhibition of respiration by endogenous nitric oxide in intact cells. Biochim Biophys Acta 1797: 557-565.
- MiPNetAnalyzer
- Hickey AJ, Renshaw GM, Speers-Roesch B, Richards JG, Wang Y, Farrell AP, Brauner CJ (2012) A radical approach to beating hypoxia: depressed free radical release from heart fibres of the hypoxia-tolerant epaulette shark (Hemiscyllum ocellatum). J Comp Physiol B 182: 91-100.
- Krumschnabel G, Eigentler A, Fasching M, Gnaiger E (2013) Combined high-resolution respirometry and fluorometry. Validation of safranin for determination of mitochondrial membrane potential. Methods Enzymol (in press).
- Gnaiger E (2012) Mitochondrial Pathways and Respiratory Control. An Introduction to OXPHOS Analysis. 3rd ed. Mitochondr Physiol Network 17.18. OROBOROS MiPNet Publications, Innsbruck: 64 pp.