On the Possibility of Real-Time Monitoring of Glucose in Cell Culture by Microdialysis Using a Fluorescent Glucose Binding Protein Sensor

• Published in: Biotechnology progress Vol. 24; no. 3; pp. 691 - 697
• Main Authors: Ge, Xudong, Rao, Govind, Tolosa, Leah
• Format: Journal Article Conference Proceeding
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.05.2008; American Chemical Society; American Institute of Chemical Engineers
• Subjects: Biological and medical sciences; Biosensing Techniques - methods; Biotechnology; Computer Systems; Culture Media - analysis; Feasibility Studies; Fundamental and applied biological sciences. Psychology; Glucose - analysis; Microdialysis - methods; Monitoring, Physiologic - methods; Protein Binding; Proteins - analysis; Proteins - chemistry; Spectrometry, Fluorescence - methods; Cell culture; Binding protein; Detector; Microdialysis; Fluorescence; Glucose; Real time; Monitoring
• ISSN: 8756-7938; 1520-6033
• DOI: 10.1021/bp070411k

Although glucose sensors with millimolar sensitivity are still the norm, there is now a developing interest in glucose sensors with micromolar sensitivity for applications in minimally invasive sampling techniques such as fast microdialysis and extraction of interstitial fluid by iontophoresis and laser poration. In this regard, the glucose binding protein (GBP) with a binding constant for glucose in the micromolar range is of particular relevance. GBP is one of the soluble binding proteins found in the periplasmic space of Gram‐negative bacteria. Because of its hinge‐like tertiary structure, glucose binding induces a large conformational change, which can be used for glucose sensing by attaching a polarity sensitive fluorescent probe to a site on the protein that is allosterically responsive to glucose binding. Correspondingly, the resulting optical biosensor has micromolar sensitivity to glucose. Because binding is reversible, the biosensor is reusable and can be stored at 4 °C for 6 months without losing its sensitivity. In this paper, we show the feasibility of using the GBP biosensor to monitor glucose in microdialysis. The effect of perfusion rate, bulk glucose concentration and temperature on microdialysis efficiency was determined. Additionally, the glucose concentrations in mammalian cell culture were monitored to demonstrate the applicability of this sensor in complex and dynamic processes over a period of time. As the sensor is sensitive to micromolar glucose, high dialysis efficiency is not required when the bulk glucose concentration is within the millimolar physiological range. Thus, a perfusion rate of 10 μL/min or faster can be used, resulting in delay times of 1 min or less.