The goal of this research application is to develop a rapid, easy to use, integrated nucleic acid test for severe acute respiratory syndrome (SARS) based on our proprietary lab-in-a-tube (Liat TM) platform. The Liat system enables the integration of sample preparation, multiplex nucleic acid amplification and real-time quantification in a single closed tube system. The recent SARS epidemic showed that more sensitive, rapid, and automated nucleic acid testing capability is needed at the local level to better diagnose patients and improve quarantine effectiveness. When completed, the proposed system will be capable of taking raw plasma or sputum samples and performing fully automated sample-to-answer SARS associated coronavirus (SCV) nucleic acid testing at the point-of-care in less than 1 hour. In meeting these requirements, the system will have broad applications in bio-defense and civilian healthcare. To enable the Phase I goals several SCV-specific minor groove binding (MGB)-TaqMan probes will be designed and validated. Viral RNA isolation using silica magneti6 beads or nucleic acid hybridization methodologies will be integrated into the lab-in-a-tube testing platform. Following probe design and RNA enrichment, the system will be validated for SCV detection sensitivity and SNP analysis in less than 1 hour from sample-to-answer using reverse transcription-polymerase chain reaction in our closed system Liar Molecular Analyzer prototype. We believe this proposed research will contribute significantly to two of NIAID's top priorities: 1) improving our nation's bio-defense capability, and 2) enhancing the diagnosis and treatment of infectious diseases. ? ? ?
