tetano
Editor, Senior Moderator
Sci Rep
. 2021 Nov 1;11(1):21385.
doi: 10.1038/s41598-021-00827-0.
A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
Jonas Schmidt[SUP] 1 2 3 [/SUP], Sandro Berghaus[SUP] 1 [/SUP], Frithjof Blessing[SUP] 1 2 [/SUP], Folker Wenzel[SUP] 2 [/SUP], Holger Herbeck[SUP] 1 [/SUP], Josef Blessing[SUP] 1 [/SUP], Peter Schierack[SUP] 3 4 [/SUP], Stefan Rödiger[SUP] 3 4 [/SUP], Dirk Roggenbuck[SUP] 5 6 [/SUP]
Affiliations
Abstract
Shortages of reverse transcriptase (RT)-polymerase chain reaction (PCR) reagents and related equipment during the COVID-19 pandemic have demonstrated the need for alternative, high-throughput methods for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mass screening in clinical diagnostic laboratories. A robust, SARS-CoV-2 RT-loop-mediated isothermal amplification (RT-LAMP) assay with high-throughput and short turnaround times in a clinical laboratory setting was established and compared to two conventional RT-PCR protocols using 323 samples of individuals with suspected SARS-CoV-2 infection. Limit of detection (LoD) and reproducibility of the isolation-free SARS-CoV-2 RT-LAMP test were determined. An almost perfect agreement (Cohen's kappa > 0.8) between the novel test and two classical RT-PCR protocols with no systematic difference (McNemar's test, P > 0.05) was observed. Sensitivity and specificity were in the range of 89.5 to 100% and 96.2 to 100% dependent on the reaction condition and the RT-PCR method used as reference. The isolation-free RT-LAMP assay showed high reproducibility (Tt intra-run coefficient of variation [CV] = 0.4%, Tt inter-run CV = 2.1%) with a LoD of 95 SARS-CoV-2 genome copies per reaction. The established SARS-CoV-2 RT-LAMP assay is a flexible and efficient alternative to conventional RT-PCR protocols, suitable for SARS-CoV-2 mass screening using existing laboratory infrastructure in clinical diagnostic laboratories.
. 2021 Nov 1;11(1):21385.
doi: 10.1038/s41598-021-00827-0.
A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test
Jonas Schmidt[SUP] 1 2 3 [/SUP], Sandro Berghaus[SUP] 1 [/SUP], Frithjof Blessing[SUP] 1 2 [/SUP], Folker Wenzel[SUP] 2 [/SUP], Holger Herbeck[SUP] 1 [/SUP], Josef Blessing[SUP] 1 [/SUP], Peter Schierack[SUP] 3 4 [/SUP], Stefan Rödiger[SUP] 3 4 [/SUP], Dirk Roggenbuck[SUP] 5 6 [/SUP]
Affiliations
- PMID: 34725400
- DOI: 10.1038/s41598-021-00827-0
Abstract
Shortages of reverse transcriptase (RT)-polymerase chain reaction (PCR) reagents and related equipment during the COVID-19 pandemic have demonstrated the need for alternative, high-throughput methods for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mass screening in clinical diagnostic laboratories. A robust, SARS-CoV-2 RT-loop-mediated isothermal amplification (RT-LAMP) assay with high-throughput and short turnaround times in a clinical laboratory setting was established and compared to two conventional RT-PCR protocols using 323 samples of individuals with suspected SARS-CoV-2 infection. Limit of detection (LoD) and reproducibility of the isolation-free SARS-CoV-2 RT-LAMP test were determined. An almost perfect agreement (Cohen's kappa > 0.8) between the novel test and two classical RT-PCR protocols with no systematic difference (McNemar's test, P > 0.05) was observed. Sensitivity and specificity were in the range of 89.5 to 100% and 96.2 to 100% dependent on the reaction condition and the RT-PCR method used as reference. The isolation-free RT-LAMP assay showed high reproducibility (Tt intra-run coefficient of variation [CV] = 0.4%, Tt inter-run CV = 2.1%) with a LoD of 95 SARS-CoV-2 genome copies per reaction. The established SARS-CoV-2 RT-LAMP assay is a flexible and efficient alternative to conventional RT-PCR protocols, suitable for SARS-CoV-2 mass screening using existing laboratory infrastructure in clinical diagnostic laboratories.