key: cord-0741092-388e574g authors: Yang, Meiting; Tang, Yidan; Qi, Lijuan; Zhang, Sicai; Liu, Yichen; Lu, Baiyang; Yu, Jiaxue; Zhu, Kun; Li, Bingling; Du, Yan title: SARS-CoV-2 Point-of-Care (POC) Diagnosis Based on Commercial Pregnancy Test Strips and a Palm-Size Microfluidic Device date: 2021-08-23 journal: Anal Chem DOI: 10.1021/acs.analchem.1c01829 sha: 2940c894dffe55ce33c67fbeb7f0d534aef951f0 doc_id: 741092 cord_uid: 388e574g [Image: see text] Coronavirus diseases such as the coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), pose serious threats. Portable and accurate nucleic acid detection is still an urgent need to achieve on-site virus screening and timely infection control. Herein, we have developed an on-site, semiautomatic detection system, aiming at simultaneously overcoming the shortcomings suffered by various commercially available assays, such as low accuracy, poor portability, instrument dependency, and labor intensity. Ultrasensitive isothermal amplification [i.e., reverse transcription loop-mediated isothermal amplification (RT-LAMP)] was applied to generate intensified SARS-CoV-2 RNA signals, which were then transduced to portable commercial pregnancy test strips (PTSs) via ultraspecific human chorionic gonadotropin (hCG)-conjugated toehold-mediated strand exchange (TMSE) probes (hCG-P). The entire detection was integrated into a four-channel, palm-size microfluidic device, named the microfluidic point-of-care (POC) diagnosis system based on the PTS (MPSP) detection system. It provides rapid, cost-effective, and sensitive detection, of which the lowest concentration of detection was 0.5 copy/μL of SARS-CoV-2 RNA, regardless of the presence of other similar viruses, even highly similar severe acute respiratory syndrome coronavirus (SARS-CoV). The successful detection of the authentic samples from different resources evaluated the practical application. The commercial PTS provides a colorimetric visible signal, which is instrument- and optimization-free. Therefore, this MPSP system can be immediately used for SARS-CoV-2 emergency detection, and it is worthy of further optimization to achieve full automation and detection for other infectious diseases. All proteins were separated by 12% SDS polyacrylamide gel electrophoresis and transferred onto PVDF membranes. The membranes were blocked with 5% (w/v) dry non-fat milk and 0.05% Tween 20 in Tris-buffered saline (TBST) for 1 h, incubated with primary antibodies at 4 °C overnight and then with HRP-conjugated secondary antibodies for 40 min at room temperature. Immunoreactive bands were visualized using ECL (Beyotime, China) and digitally captured by the Bio-Image System (DNR, Israel). Figure S1 . Structure illustration of the microfluidic portable POC device. Figure S2 . Optimization of the reaction system of LAMP using fluorescence method. Tables and captions Table S1 . Related sequences of RT-LAMP and hCG-P probes. 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