Application-oriented microfluidic LOC devices for the detection of microorganisms, toxic chemicals and serological biomarkers

Abstract

Lab-on-a-chip (LOC) technology has advanced over the past several decades. As miniaturized multiphase multistep reactors, LOCs are suitable for the implementation of complex liquid phase reactions in the field of biomedical detection. This doctoral thesis focused on the development of new LOC devices and relevant functionalities for various application cases, including microbe detection, water-safety testing, while also presenting preliminary studies on the measurement of signal molecules in blood. The application-oriented R&D (research and development) strategy was employed in the studies on a series of biomedical LOC devices. Technological challenges, e.g. optimization of on-chip quantification NASBA (nucleic acid sequence-based amplification) protocols, system design etc. were resolved in individual cases. Based on a low-cost R&D strategy, most LOCs in this doctoral work were developed for bench-top equipment as disposable components. It is worth noting that a methodological concept is also developed and summarized in this thesis, i.e., the selection of decision support mechanisms (DSMs) for biomedical LOC devices. The DSM is the supporting mechanism generating measurable signals in the LOC, and translating them into a meaningful conclusion, which can help users to make decisions in the real application field. Namely, DSMs can enable LOC-based testing and are thus a distinguished feature of application-oriented LOC devices. The author categorized the research achievements presented in this thesis into three distinctive classes, as follows:

1) Conventional DSMs commonly employed in current biomedical experimental approaches. Articles I, II and III focus on this category of DSMs. The series of LOCs in these reports were modelled on standard microtiter plates, and thus these chips are completely compatible with the microplate readers commonly utilized in biological laboratories. Article I reported the design, fabrication and validation of a disposable 43-chamber LOC device for quantitative detection of waterborne pathogens. Its principle was similar to conventional ELISA(enzyme-linked immunosorbent assay) tests for microbes, entitled X. Zhao: Application-oriented microfluidic LOC devices for the detection of microorganisms, toxic chemicals and serological biomarkers immuno-NASBA assay. A synthetic peptide and two common waterborne pathogens (Escherichia coli and rotavirus) in artificial samples were used to validate the LOC functions, which indicated that the LOC device has the potential to quantify traces of waterborne pathogens with high specificity.

Article II described the development of a LOC platform for environmental investigations into aquatic microorganisms, on the basis of quantitative NASBA (Q-NASBA). The LOC system was composed of a membrane-based sampling module, a sample preparation cassette, and a 24-channel Q-NASBA chip. The DSM of the LOC was derived from the polyurethane-foam-unit (PFU) method, which has been widely used to evaluate environmental pollution in fresh water. The multifunctional system could simplify and standardize the complicated processes of microbial detection.

Article III addressed the implementation of a 384-chamber microfluidic simulator with the incorporated functions of pathogen identification and antimicrobial susceptibility testing (AST) for personalizing the antibiotic treatment of urinary tract infections (UTIs). Its DSM adopted the diagnosis principle of conventional ATP bioluminescence assay (ATP-BLA) for living microbial cells. 2) Unconventional DSMs for contrived LOC systems. The second class of biomedical LOCs employs tailored DSMs, which are still derived from known working principles. However, the concrete mechanisms and diagnostic criteria are arbitrary in contrived functional LOCs. Article IV investigated a bioluminescent-cell-based continuous-flow device, termed a ‘living-cell chip’, with a self defining DSM to implement real-time broad-spectrum online monitoring of water quality. The prototype integrated a T-junction droplet generator, counter-flow micro-mixers, and time-delay channels. The LOC device can mix the water sample and Vibrio fischeri cell sensors into a droplet flow, and incubate the droplets in the time-delay channels before optical detection. Its DSM relies upon the relationship between the toxicant concentration in the water sample and the relative luminescence units of the bioluminescent cells in the running droplets, which is obviously different from the conventional intermittent method of ISO11384. The proposed LOC system shows great promise for an early warning system against potential toxicant chemicals in drinking water.

3) Developing prognostic/diagnosis DSMs for biomedical-database-dependent LOC systems with the aid of computational modelling. The third category relies on computational modelling within a large-scale medical/healthcare database, which is currently emerging and is not yet completely developed. Article V reported a pilot study on the design and fabrication of the LOC device for signal molecule profiling in blood. Articles VI proposed the potential roadmap and preliminary experimental approach for the construction of a human signal-molecule-profiling database (HSMPD) by the use of the former LOC device, leading to prognostic/diagnosis DSMs in the future.