Gel-entrapment of perfluorocarbons: A fluorine-19 NMR spectroscopic method for monitoring oxygen concentration in cell perfusion systems
✍ Scribed by K. A. McGovern; J. S. Schoeniger; J. P. Wehrle; C. E. Ng; J. D. Glickson
- Publisher
- John Wiley and Sons
- Year
- 1993
- Tongue
- English
- Weight
- 844 KB
- Volume
- 29
- Category
- Article
- ISSN
- 0740-3194
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✦ Synopsis
Abstract
Oxygenation is a major determinant of the physiological state of cultured cells. ^19^F NMR can be used to determine the oxygen concentration available to cells immobilized in a gel matrix by measuring the relaxation rate (1/T~1~) of perfluorocarbons (PFC) incorporated into the gel matrix. In calcium alginate gel beads without cells the relaxation rate (1/T~1~) of the trifluoromethyl group of perfluorotripropylamine (FTPA) varies linearly with oxygen concentration, with a slope of 1.26 ± 0.15 × 10^−3^ s^−1^μM^−1^ and an intercept of 0.50 ± 0.04 s^−1^. During perfusion with medium equilibrated with 95%/5% O^2^/CO^2^, changes in PFC T~1~s indicate that the average oxygen concentration was reduced from 894 ± 102 μM in the absence of cells to 476 ± 65 μM and 475 ± 50 μM in the presence of 0.7 × 10^8^ EMT6/Ro and RIF‐1 murine tumor cells per milliliter of gel, respectively. The presence of 0.2 μl of FTPA/ml of gel had no effect on the energy status of the cells as indicated by ^31^P NMR spectra.
To calculate oxygen gradients within the beads from the average PFC T~1~ of the sample, a mathematical model was used assuming that oxygen is the limiting nutrient for cell metabolism and that the cellular oxygen consumption rate is independent of oxygen concentration. Data for EMT6/RO cells were fit using experimentally determined perfusion parameters together with literature values for cell volume and oxygen consumption rate. The average PFC 1/T~1~s predicted using different literature values for volume and oxygen consumption− 1.10 ± 0.10 and 1.28 ± 0.36 s^−1^−agreed well with the experimentally measured value‐1.104 ± 0.004 s^−1^. Thus, the model is a suitable tool for calculation of oxygen consumption rates from PFC T~1~s in well‐oxygenated cell perfusion systems.