![Oxygen control – Active O2 production
• Electrolysis of aqueous solutions
• Dosing by diffusion into cell culture chamber
• Voltage / Current, Time [O2]
Cell chamber
Membrane
Electrolyte O2
Platinum electrodes
This course is funded with the support of the METOXIA project
06.10.12 under the FP7 Programme.](https://image.slidesharecdn.com/hubertflamm-121025053636-phpapp01/85/Global-local-oxygen-control-in-in-vitro-systems-18-320.jpg)
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Global & local oxygen control in in vitro systems
The Metoxia course focuses on global and local oxygen control in in vitro systems, emphasizing the importance of monitoring oxygen levels for cell cultures. It covers various strategies for oxygen regulation, including incubator designs, medium height adjustments, and active oxygen production techniques. The course is supported by the Metoxia project under the FP7 programme, and discussions on the physics of oxygen dynamics and measurement technologies are integral to the curriculum.
Global & local oxygen control in in vitro systems
- 1. METOXIA Course Global and local oxygen control in in vitro systems Hubert Flamm, IMTEK Freiburg 06.10.2012 This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 2. Learning objectives •Monitoring – The only way to get oxygen under control • Cell cultures – Physics behind oxygen distribution • Oxygen production – Smart solution with potential This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 3. Monitoring – Itsworth the effort • Which oxygen tension does the cell see? • Monitoring as control loop input • How and where can we detect pericellular oxygen levels? − Optical dip-in sensor (Fluorescence) − SCCF platform (Electrochemical) This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 4. Cell culture oxygenation + well defined environment + „clean room“ conditions - long regulation path - global regulation This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 5. Physics – Culturemodel pO2 atmosphere c0 h medium ccells cells This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 6. Physics – Partialpressure What is the partial pressure of oxygen gas above the medium? n ptotal = ∑ pi i =1 ptotal = p N 2 + pCO2 + pO2 + pH 2O John Dalton, 1766-1844 Assumption: Ideal gas pO2 = vol % O2 * patm − 60hPa This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 7. Physics – Culturemodel pO2 atmosphere c0 h medium ccells cells This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 8. Physics – Solubilityof oxygen How many oxygen disolves in the cell medium? c0 = k H * pO2 c0 = k H (T , S ) * pO2 ( patm , pH 2O ) William Henry, 1774 -1836 c0 This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 9. Physics – Culturemodel pO2 atmosphere c0 h medium ccells cells This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 10. Physics - Oxygendiffusion What happens if cells metabolise oxygen? Cells consume oxygen with rate −j pO2 atmosphere c0 h medium -j ccells cells This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 11. Physics - Oxygendiffusion What happens if cells metabolise oxygen? Diffusion flux can be described by Fick´s first law ccells − c0 jh j = −D ccells = c0 + ∆h D Adolf Fick, 1829-1901 pO2 atmosphere c0 h medium -j ccells cells This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 12. Oxygen control -Strategies jh ccells = k H (T , S ) * pO2 ( patm , p H 2O ) + D This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 13. Oxygen control –Incubator box • Feedback loop CELLS / INCUBATOR – Oxygen sensor for pericellular measurement – Software interface Sensor / Incubator • Limitations – Same global atmosphere for all cell cultures – Long control loop (long response time) This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 14. Oxygen control –Single flask supply Inside incubator: – Individual gas supply in each (critical) flask for fine-tuning – Easy to implement, but additional equipment needed – Shorter regulation loop (shorter response time) This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 15. Oxygen control –Medium height Medium height is influencing oxygen concentration at cells jh ccells = c0 + D Height could be altered: – static (if something is known about oxygen consumption) – dynamic with feedback to sensor reading • Problems with forced convection Increasing viscosity or seldom adjustments This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 16. Oxygen control –Local regulation External source counter-acting the cellular oxygen consumption pO2 atmosphere c0 h medium ccells cells j prod > 0 Steady-state: ccells = c0 + (j cells + j prod ) ∗ h D This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 17. Oxygen control –Multichamber flask • Multichamber construction • Oxygen supply across thin diaphragm + short diffusion path + rapid concentration change - premixed gas required This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 18. Oxygen control –Active O2 production • Electrolysis of aqueous solutions • Dosing by diffusion into cell culture chamber • Voltage / Current, Time [O2] Cell chamber Membrane Electrolyte O2 Platinum electrodes This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 19. Oxygen control –Active O2 production Cells • high aspect ratio PDMS • integration of micro-fluidics Isolation Glass Pt • complex geometries • plasma enhanced bonding This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 20. Oxygen control –Active O2 production This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 21. Oxygen control –Active O2 production Video This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 22. Oxygen control –Active O2 production 300 290 2 250nA (0.476 mA/cm ) 280 2 200nA (0.381 mA/cm ) oxygen / µM 2 100nA (0.190 mA/cm ) 2 270 50nA (0.095 mA/cm ) 2 20nA (0.038 mA/cm ) 260 250 0 2 4 6 8 10 time / min 350 340 330 • Linear production oxygen / µM Y=A+B*X 320 Parameter Value Failure ------------------------------------------------------------ A 248,00183 2,15675 310 B 215,74322 6,16445 ------------------------------------------------------------ • Wide operation range R SD N P 300 ----------------------------------------------------------- 0,99918 1,31363 4 8,15423E-4 ------------------------------------------------------------ 290 280 0,15 0,20 0,25 0,30 0,35 0,40 0,45 0,50 2 current density / mA/cm This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 23. Oxygen control –Active O2 production cell chamber refill 370 350 • Fast response time 330 • Predefined profiles programmable oxygen / µM 310 290 270 250 8 23 38 53 68 83 98 113 128 time / min 285 275 oxygen / µM • Longtime stability 265 • Short pulses and long ramps 255 0 20 40 60 80 100 120 time / min This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 24. Oxygen control –Active O2 production Simulation as design tool Measurement as simulation input 300 290 2 250nA (0.476 mA/cm ) 280 2 200nA (0.381 mA/cm ) oxygen / µM 2 100nA (0.190 mA/cm ) 2 270 50nA (0.095 mA/cm ) 2 20nA (0.038 mA/cm ) 260 250 0 2 4 6 8 10 time / min This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 25. Oxygen control –Active O2 production Where to measure oxygen? This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 26. Oxygen control –Active O2 production Simulation as guidance for design • Lateral oxygen profiles possible • Mimicking of reperfusion This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
- 27. Coffee break • Coffee increases open-mindedness – Coffee drinkers agree rather than others • Coffein raises sexual longing – proven for female rats • Coffee improves short-term memory • Coffee consumption increases health damages caused by smoking This course is funded with the support of the METOXIA project 06.10.12 under the FP7 Programme.
Editor's Notes
- #2 This course is funded with the support of the METOXIA project under the FP7 Programme.
- #7 Wasserdampfdruck aus Clausius-Clapeyron oder Antoine-Gleichung Dry air
- #9 Wasserdampfdruck aus Clausius-Clapeyron oder Antoine-Gleichung
- #11 c = 1.5587 mol/l/hPa * p_O2
- #12 c = 1.5587 mol/l/hPa * p_O2
- #28 Bad conscience