Capacitive Biosensor - A Tool for Ultrasensitive Analysis : Application in Clinical Analysis and Process Monitoring

University dissertation from Lund University (Media-Tryck)

Abstract: Popular Abstract in English A biosensor is an analytical device that combines a transducer with a biological sensing element. This sensor is unique and has played an important role for a broad range of applications including clinical analysis and process monitoring. Between different types of biosensors, the capacitive biosensor has recently gained a lot of interest due to its analytical performances. The sensor can detect the interaction without the need of labelling compound, thus resulting in a cheap and fast analysis. Moreover, the result is obtained in real time and, most importantly, with an ultra-sensitivity characteristic to the technique. It is possible to detect the biomolecule at concentrations far below the detection limit of any other analytical technique or type of biosensor. This thesis describes the development of a capacitive bio-sensing system. The sensor consists of a gold electrode on which the biological sensing element e.g. antibody was attached. This antibody is specific toward the substance that needs to be detected. The sensor is mounted in a flow injection system, and a continuous stream of liquid containing the target is passed over the sensing surface. The interaction of the target substance with the antibody will cause the capacitance to decrease proportional to the concentration of the target substance. The development of a capacitive biosensor in this thesis focused on different aspects. The results are summarized in 6 papers. The first study (paper I) described the use of thin gold film electrodes, fabricated by thermal evaporation, as a transducer surface. This thin film electrode can be used without the need of surface pre-treatment e.g. mechanical polishing which is required for solid gold electrode. The great success demonstrated in this study led to the mass production of disposable electrodes. Another thin film deposition called “sputtering” was also studied and the electrodes fabricated from this technique were used in this thesis. To demonstrate the use of the capacitive system, the sensors were first developed for the analysis of target substances which are important in the diagnosis of diseases. The work in paper I describes the detection of human serum albumin (HSA) from human serum sample. The capacitive biosensor results provide good correspondence with the results obtained in hospital analyses. The second application in clinical analysis is the detection of HIV-1 capsid protein, p24 antigen. The analysis of p24 antigen is of considerable diagnostic interest for monitoring HIV exposure. The analytical challenge for p24 detection is the sensitivity of an analytical device as p24 antigen is only present at very low concentrations when people are initially infected with HIV. The results of this work show the ultra-sensitivity of the capacitive biosensor technique which enables HIV-1 p24 antigen detection at very low concentrations. The detection of trace impurities in the preparation of biopharmaceutical products is required for process monitoring. Therefore, high sensitivity of analytical techniques is needed as impurities are often present in minute amounts. Paper III described the development of a capacitive biosensor for the detection of host cell protein (HCPs) impurities. Host Cells e.g. E. coli used for recombinant expression are complex systems and contain hundreds, and up to thousands, of host cell proteins. These proteins can contaminate biopharmaceutical products and failure to sufficiently remove contaminants from these products can result in reduced product efficacy or adverse responses in patients. In this study, Lactate dehydrogenase (LDH) is expressed in E. coli and the production of this enzyme was used as a model system for the study of the HCPs impurities profile. The capacitive biosensor was successfully applied in the analysis of HCPs and the sensitivity of the technique is very high. This enables the detection of HCPs even when present at very low amounts. Another application in bioprocess monitoring is the detection of protein A (paper IV) – a chromatographic column ligand used during the purification of monoclonal antibodies. The leakage of protein A into monoclonal antibody product might cause a serious problem if the antibody is used as therapeutic agent. This paper described the development of the capacitive sensor which can detect protein A down to 1.0×10-17 M. The next part of this thesis describes the further development of a capacitive transducer using a new concept to measure the capacitance of the biosensor electrode (paper V). The use of the current pulse technique was found to increase the stability of the measurement. Sensor functionalities including data acquisition, automated operating system, and miniaturization are also studied in this work. The results show a good performance of the sensor which could lead to the further development of the sensor into a commercialized product, able to compete with other analytical devices in the market. Finally, the circumstances promoting the sensitivity of the capacitive biosensor were discussed (paper VI) and some theoretical calculations were provided to support the explanations. This part also comprises examples of some results and limitations of the capacitance measurement technique.

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