FLIPP NAAT A paper based LAMP assay for point of care TB detection

Abstract

Molecular diagnostic techniques like polymerase chain reaction (PCR) revolutionized the world of healthcare. Instead of waiting for infectious agents to physically grow as in conventional culture-based tests, genetic material of pathogens could be used as a biomarker to detect their presence. Though PCR has deeply penetrated and greatly benefited the field of disease diagnosis, infrastructure required to perform PCR testing makes it unaffordable and hence inaccessible for majority of world population. The COVID-19 pandemic has strongly highlighted many such grave limitations of existing molecular diagnostic systems, which are rampant in all kinds of healthcare settings. More importantly, obstacles in timely, accurate, and affordable diagnosis of infections make infectious diseases the leading cause of mortality in low-income economies. The aim of my research has been to develop an ASSURED (affordable, sensitive, specific, user-friendly, rapid and robust, equipment free and deliverable to end users) molecular diagnostic tool for infectious disease diagnosis that fits well in the purview of WHO prescribed criteria for developing point-of-care (POC) diagnostic tools. While the diagnostic tool is a platform technology, adaptable to different infectious diseases, the first application has been demonstrated for tuberculosis (TB) diagnosis because it is the deadliest infectious disease today. The developed prototype called fluorescent isothermal paper-and-plastic nucleic acid amplification test (FLIPP-NAAT) uses loop-mediated isothermal amplification (LAMP) for DNA amplification and the only ancillary equipment required for testing is a laboratory incubator. A very low-cost imaging box for filter-free end-point fluorescence detection of amplified DNA using a cell phone has also been developed. To best of our knowledge, this work is the first demonstration of a paper-based NAAT for TB starting from gDNA as template and for testing clinical patient samples using a paper-based NAAT...