Fluorophore-encapsulated nanobeads for on-site, rapid, and sensitive lateral flow assay.

Since December 2019, the rapid and sensitive detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a priority for public health. Although the lateral flow assay (LFA) sensor has emerged as a rapid and on-site SARS-CoV-2 detection technique, the conventional approach of using gold nanoparticles for the signaling probe had limitations in increasing the sensitivity of the sensor. Herein, our newly suggested methodology to improve the performance of the LFA system could amplify the sensor signal with a facile fabrication method by concentrating fluorescent organic molecules. A large Stokes shift fluorophore (single benzene) was encapsulated into polystyrene nanobeads to enhance the fluorescence intensity of the probe for LFA sensor, which was detected on the test line with a longpass filter under ultraviolet light irradiation. This approach provides comparatively high sensitivity with the limit of detection of 1 ng mL-1 for the SARS-CoV-2 spike protein and a fast detection process, which takes less than 20 min. Furthermore, our sensor showed higher performance than gold nanoparticle-based commercial rapid diagnostics test kits in clinical tests, proving that this approach is more suitable and reliable for the sensitive and rapid detection of viruses, bacteria, and other hazardous materials.

Optimization of loop-mediated isothermal amplification (LAMP) assay for robust visualization in SARS-CoV-2 and emerging variants diagnosis.

Loop-mediated isothermal amplification (LAMP) is widely used in detection of pathogenic microorganisms including SARS-CoV-2. However, the performance of LAMP assay needs further exploration in the emerging SARS-CoV-2 variants test. Here, we design serials of primers and select an optimal set for LAMP-based on SARS-CoV-2 N gene for a robust and visual assay in SARS-CoV-2 diagnosis. The limit of detectable template reaches 10 copies of N gene per 25 µL reaction at isothermal 58℃ within 40 min. Importantly, the primers for LAMP assay locate at 12 to 213 nt of N gene, a highly conservative region, which serves as a compatible test in emerging SARS-CoV-2 variants. Comparison to a commercial qPCR assay, this LAMP assay exerts the high viability in diagnosis of 41 clinical samples. Our study optimizes an advantageous LAMP assay for colorimetric detection of SARS-CoV-2 and emerging variants, which is hopeful to be a promising test in COVID-19 surveillance.