Talk:O2k-Workshops in O2k-Network Labs

Preparation for a locally organized O2k-Workshop

Organization and programme

1A. Participants

1. How many participants are expected?
2. Is it a homogenous group of either (a) experienced users or (b) unexperienced interested participants? Or is the group mixed?
3. Is a list of participants available, indicating participants who are O2k-Users?
4. Are name tags prepared?

1B. O2k-Demo

1. For beginning users, a hands-on session on instrumental assembly and service is important, and a corresponding session should be organized in small groups.
2. For an unexperienced group, an O2k-Demo oriented workshop is perferrably conducted in a seminar room.
3. Is a demo experiment to be prepared with biological samples provided locally (tissue, cultured cells)? Usually we provide freeze-dried yeast as biological sample for a demo experiment of respiration in intact cells (see below). The demo experiment can be run in a seminar room.
4. Is a projector available for PowerPoint presentations and for connecting the laptop connected with DatLab to the O2k? Electrical outlets are required (1) for the O2k, (2) ISS, (3) laptop, (4) projector.

Instruments and accessories

2A. Which and how many OROBOROS Oxygraph-2k are locally available?

1. Specify O2k-Series number(s) of demo instrument(s).
2. Are O2k-Users invited to bring their instrument for practice and guided service? Do they bring their laptop for DatLab?
3. Are there any spare parts needed from OROBOROS INSTRUMENTS?

2B. Which accessories and consumables are locally available?

1. OroboPOS-Service Kit (necessary) including OroboPOS-Membranes and OroboPOS-Electrolyte Powder (or freshly prepared electrolyte).
2. ISS-Integrated Suction System (necessary).
3. O2k-Titration Set (some Hamilton syringes are specifically required for manual titrations).
4. HRR-Dissection Set (required if permeabilized tissue preparation is to be demonstrated).
5. PBI-Shredder HRR-Set (required if tissue homogenate should be prepared).
6. Microbalance-Set (required if tissue mass, particularly wet weight, should be determined).

2C. Which O2k-Modules are locally available?

1. TIP2k-Module (optional)
2. O2k-Fluorescence LED2-Module (recommended)
3. O2k-TPP+ and Ca2+ ISE-Module (only for special ISE workshop)
4. O2k-pH ISE-Module (only for special O2k-MultiSensor workshop)
5. O2k-NO Amp-Module and separate NO sensor (only for special O2k-MultiSensor workshop)

Chemicals and consumables

3A. OroboPOS service

1. Ammonia solution concentrated, a few ml of 25% concentrated ammonia (fresh).
2. A hood or safe place (separate room) for ammonia cleaning of the OroboPOS (ask for details).

3B. Demo O2k-Core (cell respiration)

Further details: A mitochondrial reference assay for high-resolution respirometry using freeze-dried Baker’s yeast. »MiPNet15.09. Yeast: HRR Reference Assay.

1. Na-P buffer
2. 100% Ethanol in wash bottle
3. 70% EtOH in wash bottle
4. Deionised water in wash bottle
5. 1 ml pipette
6. Paper towels
7. Gloves
8. 1 rack with four 50 ml Falcons

3C. Demo O2k-Fluorometry (cell respiration and extracellular hydrogen peroxide production)

Further details: O2k-Fluorometry: Amplex® UltraRed using freeze-dried baker’s yeast. »MiPNet18.06 Amplex-Yeast.

In addition to the above, these chemicals are required:

1. Amplex red.
2. Horseradish peroxidase.
3. H2O2 solutions for calibration (see Amplex red) - needs to be prepared fresh before the experiment!
4. Ethanol and glucose stocks for substrate titrations.
5. CCCP (uncoupler; alternatively FCCP) dissolved in ethanol or DMSO.

Information and contact

• If you have any questions, please contact Verena Laner
• We bring: 20 mg yeast (pre-weighted) and 1 full package of yeast (fresh).
• See O2k-Workshops in O2k-Network Labs for programmes and further organizational details.