A simple direct RT-LAMP SARS-CoV-2 saliva diagnostic
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Abstract
Widespread, frequent testing is essential for curbing the ongoing COVID-19 pandemic. Because its simplicity makes it ideal for widely distributed, high throughput testing, RT-LAMP provides an attractive alternative to RT-qPCR. However, most RT-LAMP protocols require the purification of RNA, a complex and low-throughput bottleneck that has often been subject to reagent supply shortages. Here, we report an optimized RT-LAMP-based SARS-CoV-2 diagnostic protocol for saliva and swab samples. In the protocol we replace RNA purification with a simple sample preparation step using a widely available chelating agent, as well as optimize key protocol parameters. When tested on clinical swab and saliva samples, this assay achieves a limit of detection of 10 5 viral genomes per ml, with sensitivity close to 90% and specificity close to 100%, and takes 45 minutes from sample collection to result, making it well suited for a COVID-19 surveillance program.
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SciScore for 10.1101/2020.11.19.20234948: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
No key resources detected.
Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:A limitation of this protocol is that the increased volume of the assay leads to a reaction cost of ∼$7. This cost can be potentially reduced by reducing the volume to the 25 µl to achieve ∼$2 per reaction, albeit with reduced …
SciScore for 10.1101/2020.11.19.20234948: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
No key resources detected.
Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:A limitation of this protocol is that the increased volume of the assay leads to a reaction cost of ∼$7. This cost can be potentially reduced by reducing the volume to the 25 µl to achieve ∼$2 per reaction, albeit with reduced sensitivity. Additionally, use of open source enzymes26,33 and pooling the samples could further reduce costs. As the volume of LAMP reaction increases, false positive results seem to increase in frequency which manifested as a few negative samples turning orange in our experiments presented here. These problems could potentially be mitigated by the inclusion of UDG/UTP in the LAMP reaction, which others have used successfully to reduce false positives26. These problems could also be reduced by running reactions with 2 different primer sets, which should increase certainty of a positive or negative result. With the current surge in Covid-19 across the US and the world this technology could prove very helpful to fill the void for rapid, affordable, easily deployable, widely distributed testing.
Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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