IDBR: Development of a Universal Spectroscopic Nanosensing System for Multiplexed Genomic and Proteomic Analysis

Johns Hopkins University, Baltimore MD

Investigators

Abstract

This award supports development of an ultrasensitive spectroscopic sensing system expected to be able to detect low-abundance (zeptomoles or less) biomolecular targets without the need for amplification. This system will be capable of multiplexed analysis of DNA, RNA, and protein on a single platform. Target amplification using PCR is currently the most effective approach for detection of low-abundance nucleic acids; however, PCR tends to induce background amplification which often undermines its accuracy. Because an amplification technique for proteins is not available, detection of low-abundance is difficult. The increasing importance of integrated genomic and proteomic analysis, e.g. correlated measurements of mRNA and protein expression, calls for a universal bioanalytical tool capable of both genomic and proteomic analysis on a single platform. The sensing system will be based on innovative integration of single-molecule detection (SMD), micro/nanofluidics, and molecular biology. The system will be comprised of a generic nanosensor capable of detecting both nucleic acids and protein targets and a 4-color microfluidic SMD spectroscope capable of multiplexed fluorescence detection of single molecules. The nanosensor will consist of functionalized quantum dots (QDs) that will give a binding-induced fluorescence resonance energy transfer (FRET) signal. When combined with the 4-color microfluidic SMD spectroscope, high-throughput detection of low-abundance targets will be possible without amplification. The capability of the device will be demonstrated through simultaneous measurements of mRNA and protein expression in a model gene expression system in single cells, and through genotyping single-nucleotide polymorphisms (SNPs) in unamplified genomic DNA. The interdisciplinary nature of the effort will provide excellent opportunities for cross-disciplinary training at the graduate and undergraduate level. After successful development of the microfluidic devices, students will be involved in a series of independent research projects in the areas of BioMEMS, micro/nanofluidics, and nanobiosensor systems. High school students are also expected to participate in some aspects of the project.