Abstract
Objective: When using high-throughput batched diagnostic platforms based on RT-PCR for SARS-CoV-2 detection, avoidance of the conventional nucleic acid extraction step can help to reduce the turnaround time and increase processivity. This approach can also spare reagents and plasticware, which have experienced a shortage during the initial waves of the pandemic, reducing the overall testing costs.
Methods: This study evaluated the performance of extraction-free protocols based on simple dilution of the specimen in sterile RNAse free water (with or without a heating step) in comparison to standard RNA extraction protocols, using two commercial kits for molecular detection of SARS-CoV-2 (Allplex™ SARS-CoV-2 assay and Allplex™ SARS-CoV-2/FluA/FluB/RSV assay) in nasopharyngeal swabs (NPS).
Results: Compared with conventional protocols, extraction-free protocols based on sample dilution without a heating step exhibited a lower analytical sensitivity: 74.0% and 82.1% with the Allplex™ SARS-CoV-2 assay (tested with 139 NPS samples) and the Allplex™ SARS-CoV-2/FluA/FluB/RSV assay (tested with 69 NPS samples), with a mean increase of Ct values of +2.04 and +1.32, respectively. Most false negative results were observed with sampled low viral load. Including a step of heat exposure did not improve but actually decreased the analytical sensitivity of the assay.
Conclusions: Results confirmed that extraction-free protocols could be a faster and cheaper approach to SARS-CoV-2 detection in NPS samples, which could improve processivity of diagnostic platforms.
Keywords: SARS-CoV-2; diagnostic platform; extraction-free method; molecular diagnostics; real-time polymerase chain reaction.
【저자키워드】 SARS-CoV-2, molecular diagnostics, real-time polymerase chain reaction, diagnostic platform, extraction-free method, 【초록키워드】 pandemic, protocol, diagnostic, RT-PCR, molecular diagnostics, Nasopharyngeal swab, polymerase chain reaction, nucleic acid, SARS-CoV-2 detection, Viral, Viral load, nasopharyngeal swabs, real-time polymerase chain reaction, molecular, platform, False negative, extraction-free method, analytical sensitivity, protocols, Ct values, Dilution, Chain Reaction, water, free water, heat exposure, reagents, avoidance, specimen, extraction, nucleic acid extraction, help, reagent, standard RNA, MOST, step, NPs, approach, polymerase chain, initial, IMPROVE, Result, tested, evaluated, false negative result, exhibited, faster, reducing, reduce, Including, 【제목키워드】 faster,