Development of a colorimetric RT-LAMP-based microfluidic device for SARS-CoV-2 detection.

Area of Science:

• Molecular Biology
• Biotechnology
• Medical Diagnostics

Background:

• Accurate and rapid diagnosis of COVID-19 is crucial for controlling SARS-CoV-2 spread.
• Current methods like RT-qPCR require specialized labs and equipment.
• Loop-mediated isothermal amplification (LAMP) offers a faster, simpler alternative under isothermal conditions.

Purpose of the Study:

• To develop a colorimetric reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay for SARS-CoV-2 detection.
• To integrate the RT-LAMP assay into a microfluidic device for enhanced usability.
• To evaluate the diagnostic performance of the developed microfluidic device.

Main Methods:

• Designed RT-LAMP primers targeting conserved SARS-CoV-2 E and RdRp genes.
• Utilized hydroxy naphthol blue (HNB) for visual colorimetric detection.
• Fabricated a polydimethylsiloxane (PDMS) microfluidic device with five reaction chambers.
• Tested RNA from nasopharyngeal swabs using the RT-LAMP microfluidic device and compared with RT-qPCR.

Main Results:

• The RT-LAMP microfluidic device demonstrated 100% specificity.
• Overall sensitivity was 75.8% for samples with Ct ≤ 35.
• Sensitivity was higher for samples with lower Ct values (higher viral load): 100% for Ct ≤ 25, 88.6% for Ct ≤ 30, and 50% for 30 < Ct ≤ 35.

Conclusions:

• The colorimetric RT-LAMP microfluidic device provides simple, rapid, and reliable SARS-CoV-2 RNA detection.
• The device's visual output and minimal equipment requirements make it ideal for point-of-care and resource-limited settings.
• Performance is comparable or superior to commonly used lateral flow tests for COVID-19 diagnostics.