Publications

Testing is pivotal for early identification of disease and subsequent infection control. Pathogens’ nucleic acid sequence can change due to naturally-occurring genetic drift or intentional modification. Because of the reliance on molecular assays for human, animal, and plant disease diagnosis, we must understand how nucleotide mutations affect test accuracy. Primers designed against original lineages of a pathogen may be less efficient at detecting variants with genetic changes in priming regions. Here, we made single- and multi-point mutations in priming regions of a model SARS-CoV-2 template that was used as input for a loop-mediated isothermal amplification (LAMP) assay. We found that many of the modifications impacted assay sensitivity, amplification speed, or both. Further research exploring mutations at every position in each of the eight priming regions should be conducted to evaluate trends and determine generalizability.

Introduction: The SARS-CoV-2 pandemic, and the subsequent limitations on standard diagnostics, has vastly expanded the user base of Reverse Transcription Loop-mediated isothermal Amplification (RT-LAMP) in fundamental research and development. RT-LAMP has also penetrated commercial markets, with emergency use authorizations for clinical diagnosis. Areas covered: This review discusses the role of RT-LAMP within the context of other technologies like RT-qPCR and rapid antigen tests, progress in sample preparation strategies to enable simplified workflow for RT-LAMP directly from clinical specimens, new challenges with primer and assay design for the evolving pandemic, prominent detection modalities including colorimetric and CRISPR-mediated methods, and translational research and commercial development of RT-LAMP for clinical applications. Expert opinion: RT-LAMP occupies a middle ground between RT-qPCR and rapid antigen tests. The simplicity approaches that of rapid antigen tests, making it suitable for point-of-care use, but the sensitivity nears that of RT-qPCR. RT-LAMP still lags RT-qPCR in fundamental understanding of the mechanism, and the interplay between sample preparation and assay performance. Industry is now beginning to address issues around scalability and usability, which could finally enable LAMP and RT-LAMP to find future widespread application as a diagnostic for other conditions, including other pathogens with pandemic potential.

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Introduction: Over the past decade, loop-mediated isothermal amplification (LAMP) technology has played an important role in molecular diagnostics. Amongst numerous nucleic acid amplification assays, LAMP stands out in terms of sample-to-answer time, sensitivity, specificity, cost, robustness, and accessibility, making it ideal for field-deployable diagnostics in resource-limited regions. Areas covered: In this review, we outline the front-end LAMP design practices for point-of-care (POC) applications, including sample handling and various signal readout methodologies. Next, we explore existing LAMP technologies that have been validated with clinical samples in the field. We summarize recent work that utilizes reverse transcription (RT) LAMP to rapidly detect SARS-CoV-2 as an alternative to standard PCR protocols. Finally, we describe challenges in translating LAMP from the benchtop to the field and opportunities for future LAMP assay development and performance reporting. Expert opinion: Despite the popularity of LAMP in the academic research community and a recent surge in interest in LAMP due to the COVID-19 pandemic, there are numerous areas for improvement in the fundamental understanding of LAMP, which are needed to elevate the field of LAMP assay development and characterization.

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