Advanced Electrokinetic-Based Micro Total Analysis System for Biothreat Detection
Universal Nucleic Acid Amplification Test (NAAT) platforms rely on the discovery that conserved and variable sequences contained in all human pathogens can be exploited for development of robust rapid diagnostic methods; however, they are still costly, labor- and resource-intensive, and lacks portability, thus precluding their routine use for for both biodefense and clinical purposes in disease surveillance, field usage, or point-of care testing.
Recent advances in molecular biology and engineering have shifted toward creation of miniaturized total analysis systems (TAS) that integrate sample preparation, genetic target amplification and detection into micro/nano-scale devices on a chip with potential performance and cost advantages over conventional macroscopic instrumentations. We have made significant progress with innovative on-chip PCR approaches, but clinical relevant resutls are ultimately reliant on efficient sample processing. Based on our experience in microfluidics, we are proposing a novel approach to sample processing based entirely on a single-operation principle electrokinetic forces, for ease of integration and fabrication to achieve superior performance. Through inter-departmental as well as cross-institutional collaborations within and between Johns Hopkins University (JHU) School of Medicine, JHU Whiting School of Engineering, and MARCE, with extensive combined expertise in translating PCR, MEMS (Microelectromechanical systems), and nanotechnologies into advanced clinical diagnostics, our research objective is to develop an micro-TAS with eletrokinetic-based sample processing with on-chip PCR for broad-based bacterial detection. Specific aims include: 1) developing the essential functional components of sample process, including cell concentration, separation, lysis, transprt, and focusing; 2) designin and validating a droplet-in-oil PCR device with QD enhanced PCR assay for Eubacterial detection with early Gram-type characterization; 3) creating and evaluating a fully integrated prototype micro-TAS containing all components of sample preparation and PCR. We will adopt an already advanced broad-based PCR assay within Program Project V of MARCE-2. Our platform will also support other Program Projects in MARCE-2 and establish utility for dual use, i.e. biodefense/emerging infections and commonly encountered clinical infections.
Jeff Wang