Flow-Chamber Hybridization Rate Enhancement for Distributed Viral Load Measuremen

DESCRIPTION (provided by applicant): Viral load measurement in resource-limited settings is a persistent area of need. Recent advances in compact, easy-to-use instrumentation with low per-test costs is promising for the development of highly portable assayers (HPAs) for HIV RNA quantitation. Lacking thermal cycling and minimally susceptible to contamination, the branched DNA (bDNA) HIV RNA sandwich nucleic acid hybridization/signal amplification assay is a good candidate for viral load measurement in resource-limited settings. However, successful development of HPA-bDNA methods and instruments requires speeding hybridization of HIV RNA to solid- phase capture probes. In the current bDNA protocol, hybridization takes place overnight; such a lengthy hybridization step is impractical for HPA viral load testing in resource-limited settings. The central hypothesis of this feasibility study is that microscale fluid mechanical processes and nonlinear diffusion effects can be leveraged to significantly accelerate flow-chamber HIV RNA hybridization-to the extent that a sufficiently large fraction of total viral RNA in a sample binds to the solid phase in less than twenty minutes. A novel method referred to as sequential delamination, which takes advantage of the microscale fluid flow laminarity and nonlinear scaling of diffusive effects, will be studied. Preliminary modeling indicates that sequential delamination-enhanced flow chambers can achieve analyte capture efficiencies approaching those of lateral flow membranes (hybridization of over 30% of HIV RNA in a 100 microliter sample with a primary hybridization step only ten minutes in duration) while maintaining the operational parameter control of fully active fluid transport. To determine feasibility and lay the groundwork for HPA-bDNA system development, prototype flow chambers will be fabricated and tested using bDNA probes and reference analytes, finite element modeling will be used for design optimization and performance parameter estimation, and system requirements will be established. Plasma viral load, determined by complex blood tests, is an indication of how sick an HIV/AIDS patient is and how well he or she is responding to treatment. Regular viral load measurement is important in caring for HIV/AIDS patients, but these tests are currently not available for everyone. This project explores new ways of making viral load measurements less expensive and more readily available to doctors and patients who live far from sophisticated medical facilities.