Difference between revisions of "MiPNet10.04 CellRespiration"
From Bioblast
| Line 1: | Line 1: | ||
{{Publication | {{Publication | ||
|title= | |title=Gnaiger E (2014) An experiment with high-resolution respirometry: Phosphorylation control in cell respiration. Mitochondr Physiol Network 10.04(07): 1-12. | ||
|info=[[Media:MiPNet10.04 CellRespiration.pdf|'''Open Access-MiPNet10.04.pdf''']]; [http://www.bioblast.at/index.php/File:MiPNet10.04_CellRespiration.pdf Versions] [[Image:O2k-Protocols.jpg|right|150px|link=http://www.oroboros.at/?o2k-protocols|O2k-Protocols contents]] | |info=[[Media:MiPNet10.04 CellRespiration.pdf|'''Open Access-MiPNet10.04.pdf''']]; [http://www.bioblast.at/index.php/File:MiPNet10.04_CellRespiration.pdf Versions] [[Image:O2k-Protocols.jpg|right|150px|link=http://www.oroboros.at/?o2k-protocols|O2k-Protocols contents]] | ||
|authors=OROBOROS | |authors=OROBOROS | ||
|year= | |year=2014 | ||
|journal=Mitochondr Physiol Network | |journal=Mitochondr Physiol Network | ||
|abstract= | |abstract=Methodological and conceptual features of highresolution respirometry are illustrated in an experiment with cultured, suspended cells in the OROBOROS | ||
Methodological and conceptual features of highresolution respirometry are illustrated in an experiment with cultured, suspended cells in the OROBOROS | |||
Oxygraph-2k (O2k). The experiment demonstrates manual [http://bioblast.at/images/e/ef/MiPNet09.12_O2k-Titrations.pdf titrations] of inhibitors, and automatic titrations of an uncoupler using the electronic Titration-Injection microPump [http://www.bioblast.at/images/8/8e/MiPNet04.05_Titration-Injection.pdf TIP2k ]. Application of the DatLab 4.3 (upgraded) software is shown for instrumental control (O2k and TIP2k), and [http://www.bioblast.at/images/2/28/MiPNet12.09_O2FluxAnalysis.pdf on-line data analysis ]. The following guideline describes the experiment in the form of a laboratory protocol, complementary to the relevant sections of the Oxygraph-2k Manual. The experiments were carried out by participants of an O2k-Course on high-resolution respirometry in April 2005 (IOC30; Schröcken, Austria). | Oxygraph-2k (O2k). The experiment demonstrates manual [http://bioblast.at/images/e/ef/MiPNet09.12_O2k-Titrations.pdf titrations] of inhibitors, and automatic titrations of an uncoupler using the electronic Titration-Injection microPump [http://www.bioblast.at/images/8/8e/MiPNet04.05_Titration-Injection.pdf TIP2k ]. Application of the DatLab 4.3 (upgraded) software is shown for instrumental control (O2k and TIP2k), and [http://www.bioblast.at/images/2/28/MiPNet12.09_O2FluxAnalysis.pdf on-line data analysis ]. The following guideline describes the experiment in the form of a laboratory protocol, complementary to the relevant sections of the Oxygraph-2k Manual. The experiments were carried out by participants of an O2k-Course on high-resolution respirometry in April 2005 (IOC30; Schröcken, Austria). | ||
:>> O2k-Protocols:[http://www.oroboros.at/?o2k-protocols Overall contents] | :>> O2k-Protocols:[http://www.oroboros.at/?o2k-protocols Overall contents] | ||
:>> 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 | |mipnetlab=AT_Innsbruck_OROBOROS | ||
}} | }} | ||
| Line 26: | Line 22: | ||
|additional=O2k-Demo | |additional=O2k-Demo | ||
}} | }} | ||
[[Image:MiPNet10.04.jpg|center|width=80px|frame|Figure 1. Respiration of 32D cells (1.1106 ml-1). Traces for the two chambers of the Oxygraph-2k are superimposed. A: Oxygen flow [pmol O2s-1106]; B: Oxygen concentration [μM] (graph layout “7 Gr1-Flux Gr2-O2 Conc”). After inhibition of ATP synthase with oligomycin, FCCP was titrated at 0.5 μM steps with the TIP2k. Each titration step is automatically marked by an event (vertical lines). After the aerobic-anoxic transition, the chambers were opened for re-oxygenation, causing a 10-min disturbance of the traces of respiratory flow. The TIP2k needles were removed before opening, and re-inserted after closing the cham.]] | |||
== Further information == | == Further information == | ||
:>> [http://www.oroboros.at/?protocols_cell_pcp Excel Templates and DatLab-Demo Files] | :>> [http://www.oroboros.at/?protocols_cell_pcp Excel Templates and DatLab-Demo Files] | ||
* [[Gnaiger 2012 MitoPathways References|References]] | * [[Gnaiger 2012 MitoPathways References|References]] | ||
Revision as of 09:42, 17 February 2014
| Gnaiger E (2014) An experiment with high-resolution respirometry: Phosphorylation control in cell respiration. Mitochondr Physiol Network 10.04(07): 1-12. |
» Open Access-MiPNet10.04.pdf; Versions
OROBOROS (2014) Mitochondr Physiol Network
Abstract: Methodological and conceptual features of highresolution respirometry are illustrated in an experiment with cultured, suspended cells in the OROBOROS Oxygraph-2k (O2k). The experiment demonstrates manual titrations of inhibitors, and automatic titrations of an uncoupler using the electronic Titration-Injection microPump TIP2k . Application of the DatLab 4.3 (upgraded) software is shown for instrumental control (O2k and TIP2k), and on-line data analysis . The following guideline describes the experiment in the form of a laboratory protocol, complementary to the relevant sections of the Oxygraph-2k Manual. The experiments were carried out by participants of an O2k-Course on high-resolution respirometry in April 2005 (IOC30; Schröcken, Austria).
- >> O2k-Protocols:Overall contents
- >> Product: OROBOROS Oxygraph-2k, O2k-Catalogue
• O2k-Network Lab: AT_Innsbruck_OROBOROS
Labels: MiParea: Respiration, Instruments;methods
Organism: Mouse
Tissue;cell: Blood cells
Preparation: Intact cells
Regulation: Coupling efficiency;uncoupling, Uncoupler Coupling state: LEAK, ROUTINE, ETS"ETS" is not in the list (LEAK, ROUTINE, OXPHOS, ET) of allowed values for the "Coupling states" property.
HRR: Oxygraph-2k, TIP2k, 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
Figure 1. Respiration of 32D cells (1.1106 ml-1). Traces for the two chambers of the Oxygraph-2k are superimposed. A: Oxygen flow [pmol O2s-1106]; B: Oxygen concentration [μM] (graph layout “7 Gr1-Flux Gr2-O2 Conc”). After inhibition of ATP synthase with oligomycin, FCCP was titrated at 0.5 μM steps with the TIP2k. Each titration step is automatically marked by an event (vertical lines). After the aerobic-anoxic transition, the chambers were opened for re-oxygenation, causing a 10-min disturbance of the traces of respiratory flow. The TIP2k needles were removed before opening, and re-inserted after closing the cham.
