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UBC Theses and Dissertations

On-chip-based biosensors and point-of-care devices for label/PCR-free detection of Cryptosporidium Luka, George


Cryptosporidium, a critical waterborne protozoan pathogen found in water resources, is one of the main causes of serious illnesses and death worldwide, costing millions of dollars annually for its detection, identification and treatments. Early diagnostic techniques for detecting Cryptosporidium were established based on the existing tools such as microscopes immunoassays, and molecular techniques in specialized laboratories. However, these techniques are associated with certain limitations such as long processing times and the requirement for expensive equipments, labelling and costly reagents. The need for on-site and real-time detection in water resources and remote areas revealed the urgent need to improve the existing techniques and develop low-cost, sensitive, selective, portable, PCR/label-free and rapid detection tools for Cryptosporidium. This led to the involvement of on-chip, biosensing and nanotechnology to develop sophisticated tools to detect Cryptosporidium. In essence, on-chip, biosensing, nanotechnology, image processing, three-dimensional (3D) printing and portable diagnostic devices have now revolutionized various research fields and proved to have a great potential for point-of-care (POC) applications. However, minimal studies have been performed to incorporate these technologies in the detection of Cryptosporidium. A simple, label/PCR-free, cost-effective and portable sensing platform for detecting Cryptosporidium is yet available. In this thesis, the state-of-art on-chip-based biosensing platforms (capacitive, electrochemical and colorimetric) and POC devices are developed for sensitive, selective, rapid, portable and label/PCR-free detection of Cryptosporidium. For the capacitive and electrochemical sensors, different sensor structures are designed, fabricated, modified and tested for their selectivity, cross-reactivity, and detection limit. For the colorimetric assay, an on-chip-based PCR/label-free colorimetric assay is developed based on the aggregation of gold nanoparticles for the detection of Cryptosporidium RNA as well as providing information about Cryptosporidium species present in the water sample. A point-of-care, potable smartphone integrated on-chip system in a 3D printed holder assembly is also developed and used for color change measurements. The detection limits for the capacitive, electrochemical and colorimetric measurements are found to be 40 cells/5µl, 20 cells/5µl, and 5 µM, respectively. These results confirm that the developed on-chip biosensing platforms and portable smartphone integrated on-chip system have great potentials for the near real-time and on-site detection of Cryptosporidium oocysts.

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