MiPNet17.14 IOC72 Schroecken AT

O2k-Workshop on High-Resolution Respirometry: O2k-Basic

Schroecken, Vorarlberg, Austria 2012 December 05 to 10

Hotel Koerbersee

This workshop is a component of the K-Regio project MitoCom Tyrol.

Contact: [email protected] [email protected]

1. Workshop materials

For those who have one (or more) O2k-instruments in the lab, I recommend to bring along the O2k-Compendium (O2k-Manual). During the Workshop, we will repeatedly refer to the O2k-Manual (Open access).

The fully updated O2k-Manual, O2k-Protocols, and O2k-References (pdf reprints) will be available on your personal OROBOROS-USB included in the workshop material. Bring your laptop or use our Notebooks during the workshop, making yourself familiar with the O2k-FileFinder while our tutors can provide immediate guidance. You can download the DatLab software and many DatLab demo files from the OROBOROS-USB, such that you can practice the use of DatLab. Although our software is known as user-friendly, DatLab is always a key part of a hands-on workshop. A comprehensive methodological overview on high-resolution respirometry (Pesta and Gnaiger 2012), is recommended for reading as a good preparation for the workshop. In addition to the literature, a random or targeted walk through the O2k-Catalogue provides lots of information on the various parts of the O2k and its modular extensions.

2. Final programme

The programme can be accessed and downloaded here: soon

We keep the programme flexible, and all theoretical parts are discussion-oriented and interactive. In addition to the practical sessions, we will cover some fundamentals of mitochondrial physiology and protocol design. If you want to refresh your memory on a few conventional bioenergetic terms, take a look at the Bioblast Glossary on respiratory states - we will take some time to discuss the rationale for an extension of bioenergetics to mitochondrial physiology.

>>> Summary of links

• O2k-Manual: http://www.oroboros.at/?O2k-Manual
• 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.
• Further reading: http://www.oroboros.at/?IOC-Reading
• O2k-Catalogue_OROBOROS
• IOC72 Programme: available soon
• Glossary:_Respiratory_states

I am looking forward to an exciting O2k-Workshop, and seeing you next week in Schroecken.

Best regards

Preliminary Programme

A: Introductory lectures and hands-on experiments for basic HRR in small working teams:

A1. General HRR topics

• O2k-assembly
• Polarographic oxygen sensor (OroboPOS) service
• POS calibration
• DatLab 5 software: principles and operation
• Titration-Injection-microPump - TIP2k.

A2. Hands-on experiments with the O2k

• Measurement of instrumental background oxygen flux, manual and TIP2k-feedback control
• Intact cells: Respirometric reference assay with yeast, phosphorylation control protocol
• Preparation of permeabilized muscle fibres
• HRR with permeabilized muscle fibres.

A3. Presentation and interpretation of oxygen flux

• Normalization of respiration per tissue or per mitochondrial markers
• Coupling control ratios
• Substrate control ratios
• Generalized flux control ratios.

B: General topics

• Overview on the O2k and DatLab: High-resolution respirometry (HRR)
• Overview on O2k-MultiSensor applications
• Introduction to fundamental elements of mitochondrial respiratory physiology
• Protocols in HRR: Design and examples of experimental protocols with intact cells, permeabilized cells and tissues, isolated mitochondria.
• Quality control - towards quality assurance
• Bioblast-Session (O2k-Publications - specific topics, MiPNet-Reference laboratories, List of respirometry terms, O2k-Catalogue, Troubleshooting, user account)

C: Special interest groups

• Oxygen kinetics: experimental examples with TIP2k-feedback control (oxystat) and data analysis,
• ADP kinetics: experimental examples with TIP2-steady state control and data analysis,
• Advanced experimental protocols

Abstract Submission

Abstracts are welcome for short presentations (hot topics in mitochondrial physiology), providing participants for an opportunity to present and discuss their work, including projects in progress, methodological details and conceptual aspects.

• Submit your Abstract
• Look at submitted Abstracts >>

Lecturers and tutors

• Erich Gnaiger, Organizer, lecturer and tutor; DSL, Medical University Innsbruck, AT; OROBOROS INSTRUMENTS, AT
• Barbara Meissner, Administrative assistant and tutor; OROBOROS INSTRUMENTS, AT

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