Isothermal Nucleic Acid Amplification System for Point-of-Care HIV Diagnosis
York House School
Floor Location : M 227 H
This project presents a rapid isothermal nucleic acid amplification system capable of performing point-of-care HIV diagnoses from crude samples. The system incorporates elements of thermal lysis, recombinase polymerase amplification (RPA), and immunochromatographic strip (ICS) endpoint detection in an attempt to create a self-contained chip that possesses the sensitivity and versatility necessary to diagnose the disease in both its acute and chronic stages. The following paper outlines a strategy that is: (1) capable of performing isothermal amplification and detection of HIV DNA and RNA; (2) equipped to diagnose viral nucleic acids using raw, unprepared samples; and (3) readily adaptable for integration into a micro-device format. Initial experimentation centered on determining the ability of the RPA assay to work in conjunction with ICS to amplify and detect purified HIV DNA and RNA sequences. The focus then progressed to diagnosing viral nucleic acids using unpurified samples containing cells, proteins, and other constituents that mimic whole blood specimens. The final stages of the project involved adapting the system for integration into a microfluidic format capable of performing the test without external manipulation or access to highly specialized equipment. The system successfully detected HIV DNA in samples containing a 4% concentration of HIV genome-encoded cells. The test also produced a robust signal when applied to the detection of HIV RNA in unpurified samples. Through virtue of its rapid, isothermal, and self-contained nature, the system described here provides proof-of-concept results to support the development of a successful point-of-care HIV nucleic-acid based test.