Mobile Phone-Based Disease Detection: Towards Single-Molecule Approaches

The last 10 years have seen a number of breakthroughs in diagnostic technologies for protein biomarker detection in patient derived samples. Cutting-edge detection strategies are pushing for quantitative detection into the sub pg/mL sensitivity regime. Technologies capable of this hold great potential in disease progression monitoring and particularly, early disease detection. A particular challenge is realising methods capable of sub-pg/mL sensitivity in a cheap, robust and accessible way. Developing technologies that can be used simply and with a minimal financial burden opens up new opportunities in how diagnostic screening approaches can be employed in resource limited settings. Low-cost and cheap platform applicable for this application exist and include the lateral flow assay. However, these assays are severely limited in their performance by a number of parameters including, in particular, how well the target binding reagents (e.g antibodies) bind their targets (e.g viral proteins) for antigen testing, or how well they facilitate the detection of host antibodies (via effective presentation of viral antigen targets) in the case of serology testing for host response. In addition, the lateral flow assay is not inherently suited for single molecule detection due to its material composition and heterogeneity. This reveals two opportunities that this project seeks to address. The first is the design and optimisation of novel affinity reagents for use in lateral flow assays towards increasing the sensitivity and reducing non-specific binding through synthetic biology and design of experiment methodologies. The second, is to implement these solutions into new device architectures bridging lateral flow assays and microfluidic assays with a view to benefiting from the advantages of both to enable single-molecule detection. The following are the overarching aims of the project, and the more specific objectives which link closely to the EPSRC/UKRI strategic priorities towards a "Healthy Nation: Transforming Healthcare" through the development of new materials, sensors and imaging modalities to support low-cost and highly sensitive rapid testing.

Aims:
1. Identify limiting role of different affinity reagents in lateral flow and rapid testing formats by synthetic biology experimental and modelling routes.
2. Establish a new rapid testing platform that combines the liquid handling benefits of lateral-flow with alternative technologies such as magnetic separation and particle tracking to enable single target detection in a low-cost format.

Objectives:
1. Establish a route for the production of multivalent protein assemblies for the surface modification of nanoparticles for diagnostics.
2. Apply published modelling tools to allow the analysis and prediction of the response of these reagents in lateral flow and understand the impact of the multivalent protein structure design.
3. Develop lateral flow assays incorporating these novel reagents and assess their performance as an early implementation.
4. Produce devices capable of sample extraction and single particle analysis using fluorescent microparticles incorporating the new affinity reagents for single molecule detection using low cost fluorescence microscopy with mobile-phone readout.