Abstract

Nucleic acid tests are crucial to the identification of genetic and infectious diseases, which afflict countless people around the world. Early detection is critical in identifying treatments for these diseases and scalable and easy-to-use diagnostic technologies are needed to this end. An injection molded centrifugal microfluidic chip and pH-based detection method were developed that permit rapid, point-of-care, multiplexed nucleic acid assays. The chips featured 15 fluidically separated reaction chambers which can handle multiple diagnostic LAMP/RT-LAMP assays simultaneously. These chambers could be filled reliably to 97.5% of their capacity, enabling repeatable assays. pH-based LAMP and RT-LAMP reactions were optimized for distinguishing alleles of CYP2C19, a gene with single nucleotide polymorphisms that correlate with drug metabolism, and respiratory viral illnesses, including SARS-CoV-2. FITC-pHEMA optical pH sensors were used to generate fluorescent readouts in minimally buffered LAMP and RT-LAMP reactions, with results obtainable in under an hour. A custom-built imaging rig was used to both incubate the filled microfluidic chips and measure fluorescence from the optical pH sensors in the reaction chambers.  By pre-drying different primer sets in the reaction chambers, multiplex diagnostics of CYP2C19 alleles and respiratory viruses could be performed. This technology should enable public health officials, medical practitioners, and even home users to better assess genetic predispositions and monitor infectious agents.