Development of Simple Methods for RNA Biomarker Extraction and Detection

Abstract

Simple and rapid methods for detecting RNA biomarkers from patient samples are especially desirable in settings with limited access to laboratory resources and trained personnel. The focus of this dissertation is on the development and evaluation of simple methods to extract and quantify RNA biomarkers from patient samples that are suitable for limited resource settings. The format that we developed for processing RNA from complex samples is based on the simple idea that magnetic beads can capture RNA on their surface and be magnetically pulled across processing solution interfaces, or surface tension valves, contained within small diameter tubing without causing the solutions to intermix. The format enables sample processing without pipetting, centrifugation, or other laboratory instrumentation. This self-contained format was demonstrated to function as a simple and effective means to extract RNA or messenger RNA (mRNA) from patient samples. The format has been shown to effectively integrate with simple RNA detection methods, including an optical method called backscattering interferometry (BSI), and an isothermal nucleic acid amplification reaction called quadruplex priming amplification (QPA). This dissertation demonstrates that the self-contained format is an effective platform for the development of a complete diagnostic device with the potential to be implemented in limited resource settings.