Fully Integrated Lab-on-a-Disc for Point-of-Care Diagnostics

Abstract

Micro total analysis system(μTAS), also called lab-on-a-chip, has received great attention over the last decade due to its capability to integrate and automate the complex bioassays on a miniaturized chip with enhanced sensitivity and faster analysis time. Despite numerous examples of previously demonstrated μTAS, the fully integrated and automated devices that can be readily used in real world application have been rare. Even if lab-scale integration could be demonstrated, it often requires complex and expensive peripherals such as pumps and microscopes, which prevent translation to point-of-care type (POCT) applications. In this thesis, fully integrated lab-on-a-disc platforms were developed for ‘sample-in and answer-out’ type biomedical applications including immunoassays and molecular diagnostics. In lab-on-a-disc systems, centrifugal force generated by spinning a motor is utilized for various unit operations for microfluidic handling such as liquid transfer, metering, mixing etc. Therefore, the total processes for biomedical diagnostics could be integrated in a small sized platform, which is ideal for POCT applications. Blood is the most common sample to reveal valuable information about the health condition such as the cause of diseases and the effects of the treatment. Lab-on-a-disc has great advantage to handle blood samples because the plasma layer could be easily separated by simple centrifugation. However, the effect of the channel shape and dimension on plasma separation rate has never been explored. In chapter 2, we have investigated the geometry effects on the blood cell separation rates and significant enhancement, as high as 8 times faster separation, was achieved by using slanted channels. Furthermore, the separation rate enhancement could be predicted with a single equation for various channel width, height, tilt angle, haematocrit, etc. The optical detection is a gold standard in immunoassays. However, the electrochemical (EC) detection has its own advantages and therefore it is the most popular detection methods used in POCT applications such as diabetes tests. In EC sensors, the peripherals are usually cheaper and more importantly, the optical-grade plastic materials are not required. However, EC detection has not been demonstrated in centrifugal microfluidic platforms. In chapter 3, a fully integrated lab-on-a-disc for electrochemical detection of protein biomarkers from whole blood was demonstrated. Not only the electrochemical detection could be made from the spinning disc but also the detection sensitivity could be significantly enhanced when the EC signal was measured during the spinning. As a result, the limit of detection (LOD) of 4.9 pg/mL for C-reactive protein (CRP) detection could be achieved with the EC measurements during the flow, which was 17 folds improvement compared to the conventional optical detection. Furthermore, the proposed technique could be used to measure the local flow rate of the liquid on a spinning disc. Finally, a fully integrated centrifugal microfluidic device for the molecular diagnostics has been developed. Starting from a large volume of sample, target pathogen enrichment, DNA extraction, polymerase chain reaction (PCR), and detection steps were fully automated on a disc. To achieve this, a novel laser-mediated non-contact heating and thermally stable reversible vaiving techniques were developed. The non-contact heating enabled stable thermocycling for PCR and the simple elastic valve could prevent the evaporation issue during the PCR. As an application, food-borne pathogen, Salmonella, detection from milk sample could be demonstrated first. Here, isothermal amplification, recombinase polymerase amplification (RPA), was used instead of PCR and the total detection could be finished within 30 min. The bare eye measurement could detect Salmonella as small as the concentration of 101 cfu/mL and 102 cfu/mL in PBS and milk, respectively. Finally, the entire process for the detection of Staphylococcus aureus was fully integrated on a single disc. It enabled sample-in and answer out type test for molecular diagnosis within 90 min. In conclusion, lab-on-a-disc platforms for fully integrated biomedical diagnostics have been developed. To be specific, flow-enhanced electrochemical detection for immunoassay and molecular diagnostics for pathogen detection were demonstrated in a centrifugal microfluidic platform. It demonstrates the versatility of the centrifugal microfluidic device as a μTAS to be used in real world applications.