New Bioanalytical Approaches, Magneto Immunoassays and SIRE Technology Based Biosensors

Detta är en avhandling från Dept. of Pure and Applied Biochemistry, Lund University

Sammanfattning: This thesis presents two new technologies, SIRE technology based biosensors and magneto immunoassays, which can be used for bioanalytical applications. One of the main problems associated with biosensor technology is the instability of the recognition element since it is of biological origin and thus, has a limited lifetime. Conventionally, this problem has been solved by immobilizing the biological recognition element directly to the transducer surface, thereby, stabilizing the biomolecule and allowing for its reuse. Biosensors based on Injection of the Recognition Element (SIRE) have tried to solve the instability problem by using fresh enzyme with every measurement. The SIRE biosensor is based on a flow injection principle where a small amount of the native enzyme is injected into an internal buffer flow. The enzyme is held in direct spatial contact with the electrochemical transducer by entrapment using a semipermeable membrane. Differential measurements are performed with the SIRE biosensor allowing for control of matrix interferences. The development of the SIRE technology, as well as some clinical and industrial applications of the sensor are presented. In particular, the detection of metabolites (glucose, hydrogen peroxide, ascorbic acid and lactate) in crude samples (whole blood, cereal, baby food and tomato paste) is demonstrated. The limit of detection for the SIRE biosensor is in the micromolar range. In the second half of the thesis, the development of a new immunoassay technology based on one of the fundamental forces in nature, magnetism is presented. The magneto immunoassay technology combines the highly selective traditional immunoassay method with a new labeling technology to provide a rapid single-step analysis. The use of superparamagnetic nanoparticles as labels to antigens in heterogeneous immunoassays is demonstrated. The feasibility of using magnetic labels in immunoassays was first investigated by employing a two-site or “sandwich” assay for the detection of Concanavalin A in-buffered samples. The subsequent development of a rapid (10 minute) one-step competitive magneto immunoassay for the detection of human albumin and C-reactive protein is presented, where the limit of detection was determined to be 6 mg/l and 9.8 mg/l, respectively. The competitive assay system was applied to clinical samples (urine) for the detection of human albumin.

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