Abstract: Today, thanks to the availability of tailor made biomolecules with the desired biological functions, separations based upon (immuno)affinity techniques are more and more common in a large field of applications. By using the high selectivity of biomolecules (antibodies, receptors, specific proteins), this technique offers the possibility of isolating compounds from complex samples with a selectivity which cannot be achieved by other chromatographic methods. In order to succeed, however, the solid phase support for the immobilization of the ligand of interest plays a prominent role. For this reason, numerous supports have already been introduced while research on new materials with additional advantages is continued. Here, a new solid phase support will be discussed for (immuno)affinity applications. This material demonstrates low non-specific adsorption acid high ligand accessibility, which enables an enhanced selectivity and capacity. Because the material is available in large quantities and exhibits superb mechanical and physical stability, selective isolations have been performed on analytical as well as preparative scale. To demonstrate the potential of this new support, several applications will be presented. Based upon immunoaffinity, two applications for the determination of oestradiol in serum respectively vitamin B12 in fermentation broth will be presented. Regarding affinity chromatography, an enzyme reactor in which the enzyme glucose oxidase is immobilized on the new material, is made for the detection of glucose by Flow Injection Analysis and electrochemical detection. Next, to isolate, identify and test components on their xeno-oestrogenic activity, an affinity column is produced in which human oestrogen receptor is covalently coupled. Several components are screened on their biological activity and the results obtained will be presented here. (C) 2001 Elsevier Science B.V. All rights reserved.

Abstract: Background: Tight regulation of blood glucose levels from patients suffering from diabetes mellitus can significantly reduce the complications associated with this disease. For this reason, elaborate research efforts have been devoted to the development of a glucose sensor for the continuous in vivo monitoring of glucose. Although the use of microdialysis as a sampling interface between the body and the biosensor is widely accepted, a major drawback of conventional microdialysis is the limited in vivo recovery. Here, ultraslow microdialysis is proposed in order to obtain (near) quantitative in vivo recoveries. To avoid, however, unacceptable long delay times, the need for a small and low dead volume measuring device was recognised. Methods: A portable lightweight measuring device for continuous in vivo monitoring of glucose in subcutaneous tissue is presented. The measuring device consists of a miniaturised flow-through biosensor, connected to a microdialysis probe and a semi-vacuum pump. The biosensor is based on the amperometric detection of hydrogen peroxide after conversion of glucose by immobilized glucose oxidase. A portable potentiostat equipped with data logging is used for detection and registration. Results: The device was validated for its accuracy, precision, linearity, sensitivity, selectivity and stability during ex vivo and in vivo experiments. The linearity was found to be up to 30 mmol/l with a limit of detection of 0.05 mmol/l. The precision, depending on the biosensor tested was found to be 2-4%. No contribution to the signal could be observed from several tested electroactive species. The accuracy was found to be well in accordance with the criteria set for methods of Self Monitoring of Blood Glucose for patients with diabetes mellitus. The biosensors could be used for up to 3 days in the continuous mode. In vivo monitoring of glucose in dialysate of subcutaneous sampled tissue during glucose tolerance tests in healthy volunteers demonstrated the potential of this measuring device. Conclusions: A portable lightweight measuring device is presented which can measure continuously glucose in vivo without excessive calibration steps. The performance characteristics determined justify the application of this measuring device for the in vivo monitoring of glucose in subcutaneous sampled interstitium of diabetic patients.

Abstract: Although biosensors are by means suitable for continuous biomedical monitoring, due to fouling and blood clotting, in vivo performance is far from optimal. For this reason, ultrafiltration, microdialysis or open tubular flow is frequently used as interface. To secure quantitative recoveries of the analyte of interest, sampling at submicrolitre level will be necessary which in turn necessitates the development of small and versatile biosensor devices. Here, a miniaturised biosensor device, which directly can be connected to various interfaces will be presented. The biosensor device consists of a pulsefree pump and a biosensor with an internal volume of 10^-20 nl. In this article, the production as well as the construction of the flow-through cell of the biosensor will be discussed. The advantages and disadvantages of several production processes will be demonstrated and a detailed protocol for the production of such a nanoliter flow-through cell will be presented. With respect to the bio-selector, several permselective membranes have been tested on their performance characteristics. Results obtained with these biosensors will be presented and discussed. Finally, a protocol based upon in situ electropolymerisation for the immobilization of the biological component was defined and several biosensors based upon this principle have been produced and tested for the monitoring of glucose respectively lactate. To demonstrate, data obtained during a variety of in vivo studies at different clinical relevant applications will be presented.