Sensor in vitro

USE OF OXYGEN SENSORS AS A SURROGATE GLUCOSE SENSOR FOR IN VIVO TESTING AND IMPORTANT ISSUES RELATED TO IN VITRO SENSOR CALIBRATION... 

The main challenges in the field of implantable sensors are the stabihty of the sensor, the selectivity of the sensor, and the biocompatibihty of the sensor. First of all, the sensor must not be rejected by the body. When implanted, the sensor should operate for a prolonged time to justify any surgical procedure necessary for the introduction of the sensor into the body. Even when these two challenges are met, the sensor has typically to deal with a very complex sample matrix, most commonly blood. In vitro sensors have to cope with the same demands in terms... 

Galameau A, Primeau M, Trudeau L-E, Michnick S. -Lactamase protein fragment complementation assays as in vivo and in vitro sensors of protein-protein interactions. Nat Biotechnol 2002 20 619-622. 

Fluorescence or Forster resonance energy transfer (FRET) is widely accepted as being one of the most useful methods to observe biochemical events in vitro and in living cells. Generally, there are two forms of FRET sensors those based on a pair of genetically encoded fluorophores, usually employing fluorescent proteins from jellyfish or corals, or those based on small molecules that make use of small organic fluorophores. 

In a similar fashion, steroids are molecules that have been investigated by disruption of FRET. The sensor is a double labeled peptide with cyclodextrin bound to one side chain. The latter keeps the fluorophores closely together by accommodating the coumarin into its cavity thereby ensuring efficient FRET. Steroids compete for the cavity of cyclodextrin and displace the coumarin reducing FRET efficiency. This model, although useful for in vitro applications, seems to be poorly selective for its application in biological samples [95],... 

The first sensor proposed for detecting gastric and oesophageal pH24, made use of two fluorophores, fluorescein and eosin, immobilised onto fibrous particles of amino-ethyl cellulose, fixed on polyester foils. Only tested in vitro, the sensor reveals a satisfactory response time of around 20 seconds. 

Hansmann D.R., Gehrich G.L., Practical perspectives on the in-vitro and in-vivo evaluation of a fiber optic blood gas sensor, Proc. SPIE 906 4 (1988). 

Grant S.A., Bettencourt K., Krulevich P., Hamilton J., Glass S.R., In vitro and in vivo measurements of fiber optic and electrochemical sensors to monitor brain tissue pH, Sensors and Actuat.B 2001 72 174. 

D. Bindra, Y. Zhang, G. Wilson, R. Sternberg, D.Trevenot, G. Reach, and D. Moatti, Design and in vitro studies of a needle-type glucose sensor for subcutaneous monitoring. Anal. Chem. 63, 1692-1696 (1991). 

In contrast to other analytical methods, ion-selective electrodes respond to an ion activity, not concentration, which makes them especially attractive for clinical applications as health disorders are usually correlated to ion activity. While most ISEs are used in vitro, the possibility to perform measurements in vivo and continuously with implanted sensors could arm a physician with a valuable diagnostic tool. In-vivo detection is still a challenge, as sensors must meet two strict requirements first, minimally perturb the in-vivo environment, which could be problematic due to injuries and inflammation often created by an implanted sensor and also due to leaching of sensing materials second, the sensor must not be susceptible to this environment, and effects of protein adsorption, cell adhesion, and extraction of lipophilic species on a sensor response must be diminished [13], Nevertheless, direct electrolyte measurements in situ in rabbit muscles and in a porcine beating heart were successfully performed with microfabricated sensor arrays [18],... 

Although a few amperometric pH sensors are reported [32], most pH electrodes are potentiometric sensors. Among various potentiometric pH sensors, conventional glass pH electrodes are widely used and the pH value measured using a glass electrode is often considered as a gold standard in the development and calibration of other novel pH sensors in vivo and in vitro [33], Other pH electrodes, such as metal/metal oxide and ISFETs have received more and more attention in recent years due to their robustness, fast response, all-solid format and capability for miniaturization. Potentiometric microelectrodes for pH measurements will be the focus of this chapter. 

Various fast redox couples such as ferrocene, ferro/ferri cyanide, and ruthenium hexamine have been used as mobile mediators. In order to be electron acceptors their standard potentials must be more positive than that of FADH2/FAD redox couple (E° = 0.05 V, at pH = 7). The requirement of mobility is, however, in conflict with the lifetime of the sensor. Because the mediator is of comparable size to the substrate, it cannot be confined to the electrode proper by, for example, a dialysis membrane. In fact, the only way this type of sensor can operate is in a sample containing a sufficient concentration of the mediator (Cass et al., 1984). Obviously, this requirement makes such sensors suitable only for in vitro applications. 

Bioassays may be the most important assays since they are often the only available tools for determining the correct tertiary structure of complex protein products and the activity of even more complex biotechnology products. Bioassays are also the most problematic assays, and the variability may be 50% for animal-based bioassays. New developments in sensor technologies may improve both the speed and accuracy of bioassays [9]. The development of hematopoietic stem cells for in vitro assays has the potential to increase both the accuracy and the speed of bioassays [10]. 

Measurement. The measurement apparatus for the in vitro test of this sensor is shown in Figure 4. The sensor was dipped into the flask containing 100 ml of phosphate buffered saline solution, pH 7.4, and the output was measured with respect to stepwise changed glucose concentrations of 0 to 2000 mg/dl under the oxygen tensions of 5 to 21%, which prepared by mixing air and N2 gas. 

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