SARS-CoV-2 detection using isothermal amplification and a rapid, inexpensive protocol for sample inactivation and purification

  1. Brian A. Rabe
  2. Constance Cepko

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Abstract

This work describes the optimization of a sample preparation and detection pipeline for a SARS-CoV-2 diagnostic test that is rapid and does not require specialized equipment. This pipeline consists of viral inactivation, rendering samples safer to work with, followed by a sensitive 30-min isothermal detection reaction with a color-based red to yellow readout. Sensitivity can be further improved using a simple and inexpensive purification protocol. This pipeline can help address the shortage of testing capacity and can be run in a variety of settings.

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  1. ScreenIT

    SciScore for 10.1101/2020.04.23.20076877: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    The loop primers (LF and LB) were designed by hand, checking for appropriate melting temperatures using SnapGene software predictions.
    SnapGene
    suggested: (SnapGene, RRID:SCR_015052)
    For other collection media, the NaOH concentration will need to be optimized to ensure the pH of the final inactivated sample falls within an acceptable range such that the sample does not, upon addition, immediately cause the LAMP reaction to turn yellow or prevent the LAMP reaction from turning yellow upon successful amplification.
    LAMP
    suggested: (LAMP, RRID:SCR_001740)
    Sample Purification with Commercial Collection Media: Viral RNA was spiked directly into clean samples of Quest Diagnostics VCM or PrimeStore MTM.
    Quest
    suggested: (QUEST, RRID:SCR_005210)

    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: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    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.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1073/pnas.2011221117
  3. ScreenIT

    SciScore for 10.1101/2020.04.23.20076877: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.Randomizationnot detected.Blindingnot detected.Power Analysisnot detected.Sex as a biological variablenot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    Optimization of a Rapid Inactivation/Stabilization and Purification Protocol The current sample collection methods used for SARS-CoV-2 testing require swabs to be placed in 2-3 ml of commercial collection media, such as Quest Diagnostics Viral Collection Media (VCM)8.
    Quest
    suggested: (QUEST, SCR_005210)
    The loop primers ( LF and LB ) were designed by hand , checking for appropriate melting temperatures using SnapGene software predictions .
    SnapGene
    suggested: (SnapGene, SCR_015052)
    For other collection media , the NaOH concentration will need to be optimized to ensure the pH of the final inactivated sample falls within an acceptable range such that the sample does not , upon addition , immediately cause the LAMP reaction to turn yellow or prevent the LAMP reaction from turning yellow upon successful amplification .
    LAMP
    suggested: (LAMP, SCR_001740)

    Results from OddPub: We did not find a statement about open data. We also did not find a statement about open code. Researchers are encouraged to share open data when possible (see Nature blog).

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.

  4. Version published to 10.1101/2020.04.23.20076877 on medRxiv