Abstract
In this study, we introduce polydimethylsiloxane (PDMS)-based microfluidic devices capable of sequential dispensing of samples into multiple reaction microchambers in a single operation to provide a fast and easy sample-to-answer platform for multiplexed genetic diagnosis of multiple viral infectious diseases. This approach utilizes the loop-mediated isothermal amplification (LAMP) method to amplify and detect specific nucleic acid (DNA/RNA) targets. We present a microfluidic flow control theory for sequential liquid dispensing phenomena, which provides design guidelines for device optimization. The device specifications, such as the possible dispensing number and maximal allowable flow rate, can be theoretically designed by optimizing the geometric dimensions of the microchannels and a pair of passive stop valves integrated into each microchamber together with the water contact angles of the materials used to fabricate the microfluidic devices. In addition, a passive stop valve with a vertical-type phaseguide structure was designed to improve device performance. We could simultaneously diagnose coronavirus disease 2019 (COVID-19) and other infectious diseases, such as severe acute respiratory syndrome (SARS), seasonal influenza A, and pandemic influenza A (H1N1) 2009. The colorimetric reverse transcription LAMP (RT-LAMP) assay suggests that the four viral infectious diseases can be detected within 30 min using a hue-based quantitative analysis, and the naked eye using our microfluidic devices.
Highlights
Coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a highly infectious disease that has rapidly spread worldwide after the first cases were registered in Wuhan, China, in late December 2019.1–5 The COVID-19 pandemic has caused unprecedented disruptions in healthcare, economic, and social systems, presenting a treacherous situation that threatens people's lives, livelihoods, and well-being
For the design and optimization of our microfluidic diagnostic devices, at least two questions must be answered: 1) how many microchambers can be dispensed?, and 2) how fast can a sample-reagent mixture be introduced into the diagnostic device? Fig. 2 shows a typical experimental result, which demonstrates that a liquid is sequentially dispensed into an array of ten microchambers in a single-row format, where water colored with blue food color (0.1% w/v) was introduced into a microchannel (W = 190.6 μm, H = 51.4 μm) with a syringe pump at a flow rate of 10 μL min−1
After the sixth microchamber was filled with water, the flow of water passed through the permanent stop valve S2 of the first chamber, as shown in Video S1.† In the experiments under different flow rate conditions, once the overflow problem occurred, water was not dispensed into any more chambers
Summary
Coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a highly infectious disease that has rapidly spread worldwide after the first cases were registered in Wuhan, China, in late December 2019.1–5 The COVID-19 pandemic has caused unprecedented disruptions in healthcare, economic, and social systems, presenting a treacherous situation that threatens people's lives, livelihoods, and well-being. To prevent the spread of COVID-19, timely and accurate diagnostic testing for SARSCoV-2 is a crucial first step in enabling active surveillance, early detection, and outbreak management. The recent advent of clustered regularly interspaced short palindromic repeats (CRISPR)-based technology has made a rapid molecular diagnostics platform for COVID-19 possible with a sensitivity and specificity comparable to those of RTqPCR.[12,13] Paper-strip-based rapid antigen tests for COVID-19 offer advantages such as shorter turnaround times (up to 30 min) and reduced costs; they are inherently less sensitive because they detect specific viral proteins rather than amplified nucleic acids.[6,7] In contrast, paper-strip-based rapid antibody tests detect immunoglobulin M (IgM) and IgG antibodies directed against SARS-CoV-2 within 15–20 min. The lateral flow immunoassay technology, utilized by these tests, is acceptable as a POCT platform for COVID-19 owing
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