Synthetic Paper for Point-of-Care Diagnostics
Capillary control, surface modifications, and their applications
Time: Fri 2020-09-11 13.00
Subject area: Biotechnology Chemical Engineering Electrical Engineering Engineering Mechanics
Doctoral student: Weijin Guo , Mikro- och nanosystemteknik, Microfluidics
Opponent: Doctor Govind V. Kaigala, IBM Research - Zurich, Switzerland
Supervisor: Professor Wouter van der Wijngaart, Mikro- och nanosystemteknik; Doctor Jonas Hansson, Mikro- och nanosystemteknik
Capillary-based platforms for diagnostics are popular for the point-of-care diagnostics market because of their low cost, easy fabrication, and easy operation. Lateral flow tests are an excellent example of capillary-based platforms for point-of-care diagnostics. However, most current lateral flow tests can only provide qualitative results or have low sensitivity. Lateral flow tests with better performance are needed. In this thesis, I tried to improve the performance of lateral flow tests from different aspects: flow rate control, surface modification, plasma separation, and immunoassay application.
Variations in sample liquid properties (viscosity and surface energy) can lead to variations in the flow rate, and therefore variations in the lateral flow test results. I developed a novel capillary pump system that can provide a constant pumping flow rate independent of liquid surface energy and viscosity. This capillary pump system works well for bodily liquids, even for blood, which has low surface energy. This capillary pump system can provide a flow rate 20.96-24.76 μL/min for all the kinds of liquid tested, which is relevant for LFTs in clinical diagnostics.
The substrate of lateral flow tests should be hydrophilic and must be easy to functionalize with protein for immunoassay applications. I developed surface modification protocols for the novel lateral flow test substrate - OSTE synthetic paper. I tested different hydrophilic treatments of synthetic paper, including PEGMA, HEMA, O2 plasma, and Tween 20 coating. All these treatment methods work well, and they provide slightly different hydrophilicity, and therefore different pumping velocities for liquid samples. They can be used for applications with different requirements on hydrophilicity. Moreover, I flowed blood plasma samples through the synthetic paper with different hydrophilic treatments and found that the OSTE synthetic paper surface retains low amounts of plasma protein (with a protein recovery rate close to 100%), which shows that synthetic paper is a good material for biological sample handling. I also developed and validated two protocols for grafting on synthetic paper: thiol-yne-biotin-streptavidin and thiol-maleimide-biotin-streptavidin.
Plasma separation is an essential step for lateral flow tests using whole blood as the sample. Usually, commercial filtration membranes for plasma separation are added to the test strips. Such membranes, however, suffer from a protein retention. I built a plasma extractor using synthetic paper. The synthetic paper underwent hydrophilic treatment and was precoated with agglutination antibody. The agglutination antibody caused local agglutination of red blood cells, while the plasma continued pumping using capillary action, thus achieving the separation. The synthetic paper had a smaller surface area to make sure less protein is retained by the surface, resulting in a higher protein recovery rate (>82%) than commercial filtration membranes (73%).
Microarray technology provides a high-throughput method to test the functionality of LFT immunoassay reagents. Nitrocellulose and glass are traditional materials for microarray platforms. I used them and synthetic paper as the substrate for a protein microarray platform, and made a systematic comparison of these three substrates with respect to the fluorescence signal. To accomplish this, I used an indirect competitive sandwich assay for the detection of the antibiotic enrofloxacin in whole milk, and fluorophore as the signal label. The experiments showed that synthetic paper could provide an overall better performance in terms of signal variance, reproducibility, limit of detection, and goodness of fit. I further investigated the influence of synthetic paper geometry design on the assay performance and chose the best design for matrix study of milk. I found the matrix effect of milk to be low, and that synthetic paper can be used for enrofloxacin detection in whole milk. The LOD of enrofloxacin detection in whole milk is 1.64 nM, which is much lower than the concentration (288.98 nM) stipulated by EU regulations.The results of the microarray platform can provide guidelines for designing lateral flow tests using synthetic paper as the substrate.
These contributions can be combined or used individually to improve the performance of lateral flow tests on reproducibility and/or sensitivity. Synthetic paper has been proved to be a good substrate for LFTs by allowing easy sample handling and immunoassay coupling, and will find wider applications in the field of quantitative LFTs.