Gel-entrapment of perfluorocarbons: a fluorine-19 NMR spectroscopic method for monitoring oxygen concentration in cell perfusion systems.
Oxygenation is a major determinant of the physiological state of cultured cells. 19F NMR can be used to determine the oxygen concentration available to cells immobilized in a gel matrix by measuring the relaxation rate (1/T1) of perfluorocarbons (PFC) incorporated into the gel matrix. In calcium alginate gel beads without cells the relaxation rate (1/T1) of the trifluoromethyl group of perfluorotripropylamine (FTPA) varies linearly with oxygen concentration, with a slope of 1.26 +/- 0.15 x 10(-3) s-1 microM-1 and an intercept of 0.50 +/- 0.04 s-1. During perfusion with medium equilibrated with 95%/5% O2/CO2, changes in PFC T1s indicate that the average oxygen concentration was reduced from 894 +/- 102 microM in the absence of cells to 476 +/- 65 microM and 475 +/- 50 microM in the presence of 0.7 x 10(8) EMT6/Ro and RIF-1 murine tumor cells per milliliter of gel, respectively. The presence of 0.2 microliters of FTPA/ml of gel had no effect on the energy status of the cells as indicated by 31P NMR spectra. To calculate oxygen gradients within the beads from the average PFC T1 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.(ABSTRACT TRUNCATED AT 250 WORDS)
McGovern
K A
KA
Department of Radiology, Johns Hopkins University School of Medicine, Baltimore, Maryland 21287.
Schoeniger
J S
JS
Wehrle
J P
JP
Ng
C E
CE
Glickson
J D
JD
D000464:Alginates; D000818:Animals; D015194:Diffusion Chambers, Culture; D005461:Fluorine; D005466:Fluorocarbons; D005782:Gels; D020723:Glucuronic Acid; D006603:Hexuronic Acids; D066298:In Vitro Techniques; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008962:Models, Theoretical; D010100:Oxygen / Q000032:analysis; D010101:Oxygen Consumption / Q000502:physiology; D014407:Tumor Cells, Cultured / Q000378:metabolism*
D000464:Alginates; D005466:Fluorocarbons; D005782:Gels; D006603:Hexuronic Acids; D005461:Fluorine; D020723:Glucuronic Acid; D010100:Oxygen