Molecular diagnostics has gained attention in the field of disease diagnosis due to its exceptional sensitivity and specificity in accurately detecting various pathogens 1,2. The capacity for rapid and accurate diagnosis is crucial role in addressing various situation, ranging from managing public health emergencies (e.g., the COVID-19 pandemic) to handling seasonal infectious diseases and foodborne pathogens 3–5. The capability to early and swiftly identify threats in public health is a significant factor in disease prevention, proactive steps, and patient management 6,7. For instance, during the COVID-19 crisis, the capacity for early detection had a substantial impact on isolation measures, treatment protocols, and ultimately patient outcomes 8,9. Similarly, rapid diagnostics in the field of food safety can prevent potential outbreaks of foodborne illnesses and enable the identification of contaminants before posing significant risks to public health 10–12. The ability to promptly identify and respond to such public health threats is crucial for curbing the spread of diseases, implementing preventive steps, and effectively managing patient outcomes 13.
Amidst these diagnostic requirements, nucleic acid amplification technologies emerge as the cornerstone of molecular diagnostics. In particular, nucleic acid isothermal amplification technology provides a promising alternative to conventional field testing 14. Isothermal amplification technology optimizes processes for point-of-care (POC) applications without thermal cycling and is regarded as an alternative to PCR 15,16. This technology utilizes the principle of amplifying nucleic acids at a constant temperature, simplifying the required instrumentation and potentially reducing overall costs and equipment complexity 17,18. Among various isothermal amplification methods, Loop-mediated isothermal amplification (LAMP) is one of the preferred technologies 19. It demonstrates high sensitivity 20, specificity 21, resilience to sample impurities 22, and versatility in handling a diverse range of clinical and non-clinical samples 23, making it a suitable technology for point-of-care testing. The integration of easy-to-use platform utilizing LAMP is recognized as a significant advancement in this field 24–26. Paper-based microfluidics has been accepted as ideal POC platform due to its inherent user-friendly design and suitable property for isothermal amplification 27. These systems utilize the unique capillary action of paper to automate fluid flow, enabling the integration of automatic and power-independent analytical methods 28,29. However, despite these strides, challenges persist in developing a platform that effectively combines the convenience of paper fluidics with a reliable and accessible power source and presents an interface that is intuitive for end-users 30,31.
In this study, we developed a Lab-on-Paper (LOP) device, a rapid and highly sensitive molecular testing platform, based on LAMP and a paper strip. The developed platform consists of a USB-powered simple electric circuit and an engineered paper strip, allowing for the visual detection of LAMP amplicon within 30 minutes by the naked eye. The LOP device automates the entire process, encompassing the LAMP reaction with biotin and FAM-labeled primer on a paper strip, by thermally managing with a wax valve to transport the LAMP amplicon from the reaction zone to the detection zone on the paper strip, and conducting visual detection using anti-FAM antibody-conjugated gold nanoparticles and immobilized streptavidin in the detection zone. The LOP platform has several advantages: i) a USB-powered portable device, ii) an automated flow-controllable wax-fabricated paper strip operated by a programmed heater without intermediate steps, and iii) a low-cost and mass-producible diagnostic platform based on disposable technical paper strips. Using the LOP device, we have successfully detected Vibrio vulnificus at as low as 120 CFU/reaction and Salmonella Typhimurium at as low as 60 CFU/reaction, and those results are comparable with those from bench-top level equipment targeting the same pathogens. The detailed performance validation of the LOP platform, as outlined in the results section of this study, underscores its applicability to not only food safety testing but also various diagnostic fields where rapid, accurate, and cost-effective diagnostics are crucial. Our research work provides a comprehensive overview from conceptualization to empirical validation of the LOP platform, underscoring it as a robust platform that could be widely adopted in point-of-care testing and global health diagnostics.