Fusogenic-Reactor-Based Lateral Flow Assay for Extraction-free, Multiplexed Viral RNA Detection

Descripción del proyecto

Prueba multianalito casera para la detección de virus

Los ensayos multianalito permiten detectar simultáneamente varios virus, como el de la gripe A y el coronavirus del síndrome respiratorio agudo grave de tipo 2, y tienen una gran importancia para una vigilancia sanitaria eficaz. Las pruebas diagnósticas de virus para uso doméstico carecen de precisión, mientras que las pruebas basadas en la reacción en cadena de la polimerasa (PCR) son laboriosas y requieren recursos especializados. En el proyecto FusoLipo-LFA, que cuenta con el apoyo de las acciones Marie Skłodowska-Curie, se desarrollará una nueva prueba basada en vesículas lipídicas que imitan a las células y pueden fusionarse con virus. Estas vesículas contienen herramientas moleculares especiales capaces de detectar ARN vírico y amplificar la señal, lo que facilita la detección con tan solo unas pocas partículas víricas. Este planteamiento evita los complejos procesos de extracción y ofrece pruebas caseras rápidas, precisas y fáciles de usar la detección de múltiples virus.

Objetivo

In this fellowship, I will leverage recent advances in membrane-fusion-based biosensor technologies to develop an extraction-free, multiplexed lateral flow assay (LFA) for rapid, home-based viral RNA detection. The COVID-19 pandemic exposed critical limitations in virus diagnostics, with antigen-based kits lacking sensitivity and specificity, while RT-qPCR tests, though accurate, are slow, resource-intensive, and impractical for widespread deployment—highlighting the urgent need for field-deployable molecular assays that deliver reliable results. Given the coexistence of multiple respiratory viruses, multiplexed detection is also crucial. This project aims to combine my expertise in membrane-fusion-based biosensor design with Professor Dame Molly Stevens' (University of Oxford) expertise in LFA-based biosensors to create a multiplex LFA system for viral RNA detection using membrane fusion approach. The system will first employ cell-mimetic fusogenic lipid vesicles (FusoLipo) encapsulating CRISPR toolkits, facilitating viral membrane fusion, RNA capture, and CRISPR-mediated detection. By utilizing two types of FusoLipo, the platform can simultaneously detect influenza A (H1N1) and SARS-CoV-2 without the need for complex RNA extraction, minimizing contamination risks. Once the vesicle’s cargo is applied to the LFA, catalytic nanoparticles (Nanozyme) will amplify the signal at the detection bands, increasing sensitivity. This platform ultimately provides a accurate, and user-friendly solution for home-based multiplexed viral RNA detection. Through this fellowship, I will gain access to specialized expertise and cutting-edge facilities in lipid vesicle formulation, molecular technology, LFA development, and inorganic materials synthesis in the Stevens group, alongside advanced instrumentation to characterize vesicles at the single-particle level and further study virus-FusoLipo membrane fusion, significantly advancing my capabilities in diagnostic biosensor innovation.