Enhanced Point-of-Care SARS-CoV-2 Detection: Integrating RT-LAMP with Microscanning

Abstract

The COVID-19 pandemic has highlighted the urgent need for rapid and accurate diagnostic methods for various infectious diseases, including SARS-CoV-2. Traditional RT-PCR methods, while highly sensitive and specific, require complex equipment and skilled personnel. In response, we developed an integrated RT-LAMP-MS assay, which combines rapid reverse transcription loop-mediated isothermal amplification (RT-LAMP) with microscanning (MS) technology for detecting SARS-CoV-2. The assay uses magnesium pyrophosphate formed during LAMP amplification as a visual marker, allowing direct observation via microscopy without the need for additional chemical indicators or probes. For the SARS-CoV-2/IC RT-LAMP-MS assay, the sample-LAMP reagent mixture was added to a microchip with SARS-CoV-2 primers and internal controls, then incubated at 62 °C for 30 min in a heat block, followed by amplification analysis using a microscanner. In clinical tests, the RT-LAMP-MS assay showed 99% sensitivity and 100% specificity, which is identical to the RT-LAMP results and comparable to the commercial AllplexTM SARS-CoV-2 assay results. Additionally, the limit of detection (LOD) was determined to be 10−1 PFU mL−1 (dynamic range: 103~10−1 PFU mL−1). The assay delivers results in 30 min, uses low-cost equipment, and demonstrates 100% reproducibility in repeated tests, making it suitable for point-of-care use in resource-limited settings.

Keywords

• RT-LAMP
• SARS-CoV-2
• visual detection
• magnesium pyrophosphate
• point-of-care testing