Ultra-sensitive nanozyme-based chemiluminescence paper test for rapid diagnosis of SARS-CoV-2 infection

  1. Dan Liu
  2. Chenhui Ju
  3. Chao Han
  4. Rui Shi
  5. Xuehui Chen
  6. Demin Duan
  7. Jinghua Yan
  8. Xiyun Yan

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Abstract

The recently emerged coronavirus disease COVID-19 has now evolved into a global pandemic. Early detection is crucial for its effective control. Nucleic acid testing for viral pathogen and serological testing for host antibodies are playing important roles in current COVID-19 diagnosis. However, while nucleic acid testing is complicated, facility-restricted and time-consuming, antibody testing may result in high rates of false-negative diagnoses, especially during the early stages of viral infection. Thus, a more rapid and reliable test for both early COVID-19 diagnosis and whole-population screening is urgently needed. Here, we developed a novel nanozyme-based chemiluminescence paper assay for rapid and high-sensitive testing of SARS-CoV-2 spike antigen. Our paper test uses a newly established peroxidase-mimic Co-Fe@hemin nanozyme instead of natural HRP that catalytically amplifies the chemiluminescent signal, allowing for target concentrations to be as low as 0.1 ng/ml. Furthermore, our nanozyme-based chemiluminescence test exhibits a linear range that is 32-fold wider compared to ELISA tests. Importantly, testing is completed in less than 16 min, compared to 1-2 h required for ELISA or nucleic acid tests. Critically, signal detection is feasible using a smartphone camera. Ingredients for our test are simple and readily available, rendering overall cost considerably lower than those used in current diagnoses. In conclusion, our novel test provides a high-sensitive, point-of-care testing (POCT) approach for SARS-CoV-2 antigen detection, which should greatly increase current early screening capacities for suspected infections, and considerably lower demand for national healthcare resources.

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  1. ScreenIT

    SciScore for 10.1101/2020.06.05.131748: (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.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Fiberglass pads, absorbent pads and plastic boards were purchased from Shanghai Kinbio Tech. Co. Ltd. Ab1, Ab2, TP52, RP01 antibodies against SARS-CoV-2 spike antigen were provided by Academy of Military Medical Sciences and Sino Biological Inc. (Beijing, China).
    TP52
    suggested: None
    Anti-Spike antibodies were generated by sorting single memory B cells as previously reported19.
    Anti-Spike
    suggested: None
    , anti-CD19, anti-CD38, anti-CD27 and anti-His antibodies.
    anti-CD19
    suggested: None
    anti-CD38,
    suggested: None
    anti-CD27
    suggested: None
    anti-His
    suggested: None
    Fabrication of nanozyme-based chemiluminescence test paper: Anti-S-RBD antibody pairs were screening using ELISA method and nanozyme-based colorimetric strips15.
    Anti-S-RBD
    suggested: None
    The paired capture antibody of S-RBD (S-cAb) was immobilized on a nitrocellulose membrane at 1 mg/ml to form the test line (T-line), as well as the anti-human IgG antibody as the control line (C-line).
    anti-human IgG
    suggested: None
    Nanozyme probes uncombined with antigen bound with the control IgG antibody at C-line.
    control IgG
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    A cDNA sequence encoding S-RBD was expressed containing both N-terminal natural signal peptide as well as a 6×His tag at the C-terminus, after transfection into HEK293 cells.
    HEK293
    suggested: CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045)
    Rapid testing of pseudo-SARS-CoV-2: The pseudo-SARS-CoV-2 were packaged by transfecting 293T cells with a HIV-1-based retroviral vector coding SARS-CoV-2 spike S1 protein and luciferase reporter gene.
    293T
    suggested: None
    Software and Algorithms
    SentencesResources
    The antibody binding affinity for S-RBD was evaluated by BIAcore 8K.
    BIAcore
    suggested: (Biacore T100 System, RRID:SCR_019679)

    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 10.1101/2020.06.05.131748 on bioRxiv