Artikel
Anwendung eines laser-basierten Sensors zur Echtzeit-Sauerstoffmessung in drei-dimensionalen Gewebekulturen unter Perfusion
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Veröffentlicht: | 10. September 2012 |
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Gliederung
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Fragestellung: Die Kultivierung von Gewebekonstrukten im Tissue Engineering in klinisch-relevanten Grössen stellt eine Herausforderung an die Versorgung der Zellen mit Sauerstoff und Nährstoffen dar. Die Sauerstoffkonzentration innerhalb von dreidimensionalen Konstrukten bestimmt Zellvitalität und -metabolismus. Derzeitige Technologien zum Sauerstoffmonitoring in Bioreaktoren beinhalten fiberoptische Sensoren zur direkten Insertion in eine Matrix oder indirekte Messverfahren durch Differentialsensoren.
Methods: A real-time two-frequency phase-modulation technique to determine phosphorescent lifetime of oxygen-sensitive probes encapsulated in microbeads was applied towards mammalian static and dynamic cell culture systems. The oxygen sensor system consisted of an optical device with phosphorescent microbeads and a laser light source, a minaturized electronical device with implemented lock-in-amplifyer and frequency generator and software for data collection and analysis. Cylindrical collagen sponge-like scaffolds with microbeads were seeded with adipose-derived mesenchymal stem cells and cultured statically and dynamically in a laminar flow bioreactor up to three weeks.
Results: Signal detection was reliable at a depth of 10mm inside the collagen scaffold. MSC demonstrated a gradual decrease in oxygen concentration in the scaffold centre from 21% down to 10%.
Conclusions: The multifrequency phase-modulation technique allows measurement of phase shift of luminescent signals at two different frequencies and therefore can overcome autofluorescent signals from the tissue itself. The technique is applicable for continuous oxygen monitoring in static and dynamic culture of 3D tissue.