Long-term continuous monitoring of dissolved oxygen in cell culture medium for perfused bioreactors using optical oxygen sensors.
For long-term growth of mammalian cells in perfused bioreactors, it is essential to monitor the concentration of dissolved oxygen (DO) present in the culture medium to ascertain the health of the cells. An optical oxygen sensor based on dynamic fluorescent quenching was developed for long-term continuous measurement of DO for NASA-designed rotating perfused bioreactors. Tris(4,7-diphenyl-1,10-phenanthroline) ruthenium(II) chloride is employed as the fluorescent dye indicator. A pulsed, blue LED was chosen as the excitation light source. The sensor can be sterilized using an autoclave. The sensors were tested in a perfused rotating bioreactor supporting a BHK-21 (baby hamster kidney) cell culture over one 28-day, one 43-day, and one 180-day cell runs. The sensors were initially calibrated in sterile phosphate-buffered saline (PBS) against a blood-gas analyzer (BGA), and then used continuously during the entire cell culture without recalibration. In the 180-day cell run, two oxygen sensors were employed; one interfaced at the outlet of the bioreactor and the other at the inlet of the bioreactor. The DO concentrations determined by both sensors were compared with those sampled and measured regularly with the BGA reference. The sensor outputs were found to correlate well with the BGA data throughout the experiment using a single calibration, where the DO of the culture medium varied between 25 and 60 mm Hg at the bioreactor outlet and 80-116 mm Hg at the bioreactor inlet. During all 180 days of culture, the precision and the bias were +/-5.1 mm Hg and -3.8 mm Hg at the bioreactor outlet, and +/- 19 mm Hg and -18 mm Hg at inlet. The sensor dynamic range is between 0 and 200 mm Hg and the response time is less than 1 minute. The resolution of the sensor is 0.1 mm Hg at 50 mm Hg, and 0.25 mm Hg at 130 mm Hg.
Gao
Frank G
FG
Wyle Laboratories, Life Sciences, Systems and Services, 1290 Hercules Drive, Suite 120, Mail Stop BT-37, Houston, Texas 77058, USA.
Jeevarajan
Antony S
AS
Anderson
Melody M
MM
D000818:Animals; D016477:Artifacts; D019149:Bioreactors; D015374:Biosensing Techniques / Q000295:instrumentation; D001784:Blood Gas Analysis / Q000379:methods; D002478:Cells, Cultured; D006224:Cricetinae; D003470:Culture Media / Q000737:chemistry*; D046508:Culture Techniques / Q000295:instrumentation / Q000379:methods*; D005456:Fluorescent Dyes; D055095:Optics and Photonics; D009942:Organometallic Compounds / Q000737:chemistry; D010100:Oxygen / Q000032:analysis*; D010477:Perfusion; D010618:Phenanthrolines / Q000737:chemistry
D003470:Culture Media; D005456:Fluorescent Dyes; D009942:Organometallic Compounds; D010618:Phenanthrolines; C041132:tris(4,7-diphenyl-1,10-phenanthroline)ruthenium (II); D010100:Oxygen