Biosensor systems for blood lactate monitoring
Abstract: Enzyme based biosensor systems for ex vivo blood lactate monitoring were developed. The first approach utilised an immobilised enzyme reactor incorporated in a flow system and the main objectives were integration of sampling, sample handling, analysis, control and calibration. The flow system comprised a double lumen catheter for continuous sampling and heparinisation of blood, a dialysis unit for separation of macromolecules from the sample and an enzyme reactor containing lactate oxidase co-immobilised with catalase onto controlled pore glass for conversion of lactate. Detection was based on registration of the oxygen depletion taking place during substrate conversion and a differential measurement mode was employed to eliminate disturbances due to fluctuations in background oxygen levels. Control and calibration of the integrated system was possible by means of a microprocessor or computer. Operational stability was investigated using in vitro blood samples and during ex vivo monitoring in humans during physical exercise as well as in patients during open-heart surgery. Improved control and evaluation of sensor drift during long-term monitoring was realised through the development of a novel calibration method, based on the internal standard principle. The flow system could be calibrated in 5.5 min without interruption of the sampling step by addition of an internal standard at the tip of the double lumen catheter. Accordingly, not only the analytical step but the system in its entirety was calibrated. The second approach was based on enzyme electrodes. Lactate oxidase was immobilised between polymeric membranes and combined with an electrode system based on classical amperometric detection of hydrogen peroxide. The aim was to develop a sensor system for direct measurement in whole blood, thus eliminating the need for sample pre-treatment. Blood contacting membranes with unique haemocompatibility properties were produced through modification of poly(vinyl chloride), PVC, membranes with non-ionic surfactant. Sensor linear range could be adjusted by varying the amount of added surfactant and thereby affecting the permeability of the external PVC membranes. A range of different polymeric materials were evaluated for use as permselective internal membranes. Selectivity against common interfering compounds such as ascorbate, urate and acetaminophen as well as adequate permeability to hydrogen peroxide were the main criteria. Functional haemocompatibility was demonstrated for enzyme electrodes evaluated in whole blood and such electrodes were later incorporated in a flow system for blood lactate measurement under continuous flow conditions. Without the need for a dialysis step, the lag time for transport of sample could be decreased from 5 min to 1-2 min thereby approaching real-time monitoring.
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