Abstract

A segmented multiplex continuous-flow polymeric RT-PCR microfluidic system with a coordinate-based quantitative detection for point-of-care gene expression analysis

Geok Soon Lim, PhD Student, Nanyang Technological University

The need for rapid quantification of multiple genes of interest (GOIs) on a low-cost, disposable and easy-to-use microfluidic platform is critical in point-of-care testing (POCT) gene expression analysis applications, including companion diagnostics and personalized healthcare. Currently, most continuous-flow PCR systems are hindered by a lack of multiplexing capabilities, requiring large equipment footprint and complex detection setup, rendering them largely incompatible with POCT. In this presentation, we describe a rapid segmented continuous-flow RT-PCR microfluidic system with an adopted coordinate-based quantitative detection methodology. We show here, for the first time, a primer preloading procedure in a continuous-flow PCR system to simplify off-chip handling procedures and to allow for ease-of-integration with potential upstream modules. We validate our microfluidic system in a POCT application by performing a duplex gene expression analysis on mice tissue samples after a cyclophosphamide administration. The entire RT-PCR process is completed in <20mins – 3.9-fold faster than a conventional thermal cycler. In short, we demonstrate the efficacy of our multiplex continuous-flow RT-PCR microfluidic system, and its applicability in rapid POCT hepatotoxicity assessment.