Evaluating an extraction-free sample preparation method for multiplex detection of SARS-Cov-2, influenza A/B, and RSV with implementation on a microfluidic chip

Fatemeh Ahmadi; Fatemeh Zahra Zanganeh; Iman Amani Tehrani; Saeed Shoaee; Hamzeh Choobin; Ali Bozorg; Mojtaba Taghipoor

doi:10.1016/j.diagmicrobio.2024.116325

Evaluating an extraction-free sample preparation method for multiplex detection of SARS-Cov-2, influenza A/B, and RSV with implementation on a microfluidic chip

Diagn Microbiol Infect Dis. 2024 Apr 25;109(3):116325. doi: 10.1016/j.diagmicrobio.2024.116325. Online ahead of print.

Authors

Fatemeh Ahmadi¹, Fatemeh Zahra Zanganeh¹, Iman Amani Tehrani², Saeed Shoaee², Hamzeh Choobin³, Ali Bozorg¹, Mojtaba Taghipoor⁴

Affiliations

¹ Department of Biotechnology, College of Science, University of Tehran, Tehran, I.R., Iran.
² Micro Nano System Laboratory (MNSL), Department of Mechanical Engineering, Sharif University of Technology, Tehran, Iran.
³ Department of Virology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
⁴ Micro Nano System Laboratory (MNSL), Department of Mechanical Engineering, Sharif University of Technology, Tehran, Iran. Electronic address: mtaghipoor@sharif.ir.

PMID: 38688146
DOI: 10.1016/j.diagmicrobio.2024.116325

Abstract

Following the relaxation of COVID-19 restrictions, other respiratory viruses such as influenza and respiratory syncytial virus (RSV), whose transmission were decreased due to COVID-19 precautions, are rising again. Because of similar clinical features and reported co-infections, multiplex detection of SARS-CoV-2, influenza A/B, and RSV is required to use specific treatments. This study assessed an extraction-free sample preparation (heat treatment at 95°C for 3 minutes) for multiplex detection using rRT-PCR. Despite an observed Ct-delay (∆Ct) averageing 1.26 compared to the standard method, an acceptable total sensitivity of 92 % and a negative predictive value (NPV) of 96 % were obtained. Moreover, Implementation on a microfluidic chip demonstrated efficiency, maintaining an excellent correlation (R²=0.983) with the standard method. Combining this extraction-free procedure with rRT-PCR on a microfluidic chip seems promising, because it simplifies the design and reduces the cost and complexity of the integrated assay for multiplex detection of SARS-CoV-2, influenza A/B, and RSV.

Keywords: Lab-on-a-chip; Molecular diagnostics; Point-of-care tests; Real-time RT-PCR; Respiratory infections.