Significance: An important limitation of current assays for the detection of SARS-CoV-2 stems from their reliance on time-consuming, labor-intensive, and laboratory-based protocols for viral isolation, lysis, and removal of inhibiting materials. While RT-PCR remains the gold standard for performing clinical diagnostics to amplify the RNA sequences, there is an urgent need for alternative testing platforms that are rapid, accurate, simple, and portable. Here, we demonstrate isothermal RT-LAMP nucleic acid-based detection of SARS-CoV-2 with an additively manufactured cartridge and a smartphone-based instrument for testing that can be performed at the point of sample collection.

Point-of-care (POC) detection technologies that enable decentralized, rapid, sensitive, low-cost diagnostics of COVID-19 infection are urgently needed around the world. With many technologies approved for commercialization in the past 10 months, the field of COVID-19 POC diagnostics is rapidly evolving. In this Perspective, we analyze the current state of POC technologies for the diagnosis and monitoring of COVID-19 infection and discuss future challenges in COVID-19 diagnostics. As the COVID-19 pandemic becomes endemic, the advances gained during this past year will likely also be utilized for future prediction of emerging outbreaks and pandemics.

Rapid and accurate detection of DNA from disease-causing pathogens is essential for controlling the spread of infections and administering timely treatments. While traditional molecular diagnostics techniques like PCR are highly sensitive, they include nucleic acid amplification and many need to be performed in centralized laboratories, limiting their utility in point-of-care settings. Recent advances in CRISPR-based diagnostics (CRISPR-Dx) have demonstrated the potential for highly specific molecular detection, but the sensitivity is often constrained by the slow trans-cleavage activity of Cas enzymes, necessitating preamplification of target nucleic acids. In this study, we present a CRISPR-Cascade assay that overcomes these limitations by integrating a positive feedback loop that enables nucleic acid amplification-free detection of pathogenic DNA at atto-molar levels and achieves a signal-to-noise ratio greater than 1.3 within just 10 min. The versatility of the assay is demonstrated through the detection of bloodstream infection pathogens, including Methicillin-Sensitive Staphylococcus aureus (MSSA), Methicillin-Resistant Staphylococcus aureus (MRSA), Escherichia coli , and Hepatitis B Virus (HBV) spiked in whole blood samples. Additionally, we introduce a multiplexing OR-function logic gate, further enhancing the potential of the CRISPR-Cascade assay for rapid and accurate diagnostics in clinical settings. Our findings highlight the ability of the CRISPR-Cascade assay to provide highly sensitive and specific molecular detection, paving the way for advanced applications in point-of-care diagnostics and beyond.