A lab-on-a-chip platform for integrated extraction and detection of SARS-CoV-2 RNA in resource-limited settings

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the unprecedented global pandemic of coronavirus disease-2019 (COVID-19). Efforts are needed to develop rapid and accurate diagnostic tools for extensive testing allowing for effective containment of the infection, via timely identification and isolation of SARS-CoV-2 carriers. Current gold standard nucleic acid tests require many separate steps that need trained personnel to operate specialist instrumentation in laboratory environments, hampering turnaround time and test accessibility, especially in low-resource settings. We devised an integrated on-chip platform coupling RNA extraction based on immiscible filtration assisted by surface tension (IFAST), with RNA amplification and detection via colorimetric reverse-transcription loop mediated isothermal amplification (RT-LAMP), using two sets of primers targeting open reading frame1a (ORF1a) and nucleoprotein (N) genes of SARS-CoV-2. Results were identified visually, with a colour change from pink to yellow indicating positive amplification, and further confirmed by DNA gel electrophoresis. The specificity of the assay was tested against HCoV-OC43 and H1N1 RNAs. The assay based on use of gene N primers was 100% specific to SARS-CoV-2 with no cross-reactivity to HCoV-OC43 nor H1N1. Proof-of-concept studies on water and artificial sputum containing genomic SARS-CoV-2 RNA showed our IFAST RT-LAMP device to be capable of extracting and detecting 470 SARS-CoV-2 copies mL -1 within 1 h (from sample-in to answer-out). IFAST RT-LAMP is a simple-to-use, integrated, rapid and accurate COVID-19 diagnostic platform, which could provide an attractive means for extensive screening of SARS-CoV-2 infections at point-of-care, especially in resource-constrained settings.

Microfluidics-Based Point-of-Care Testing (POCT) Devices in Dealing with Waves of COVID-19 Pandemic: The Emerging Solution

“…Optical-based detection techniques for detection of respiratory viruses. Reproduced with permission from ref ( 130 ). Copyright 2021 Elsevier.…”

LAMP-Based Point-of-Care Biosensors for Rapid Pathogen Detection

“…Recently, microfluidic devices are also integrated with RT-LAMP for the detection of the highly infectious SARS-CoV-2 virus. For example, Mateos et al [ 126 ] devised a PMMA-made, integrated microfluidic chip for magnetic bead-based RNA extraction and RT-LAMP reaction targeting ORF1a and N gene of SARS-CoV-2. The microchip contains a large chamber for sample loading, followed by seven wash chambers and a detection chamber.…”

Sample Preparation for Lab-on-a-Chip Systems in Molecular Diagnosis: A Review

“…The emergence of the SARS-CoV-2 in the past year has rushed many scientists to come up with a rapid and easy detection platform for the virus. Rodriguez-Mateos recently reported a device that successfully detects the virus within an hour, the efficient detection occurs due to the use of oligo (dT)-coated magnetic beads extraction of viral RNA for a LAMP assay, all in a single apparatus . This was performed in a PMMA device where the extraction method was based on immiscible filtration assisted by surface tension (IFAST).…”