Rapid and quantitative detection of COVID-19 markers in micro-liter sized samples

  1. Xiaotian Tan
  2. Cory Lin
  3. Jie Zhang
  4. Maung Kyaw Khaing Oo
  5. Xudong Fan

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Abstract

COVID-19 pandemic has caused tens of thousands of deaths and is now a severe threat to global health. Clinical practice has demonstrated that the SARS-CoV-2 S1 specific antibodies and viral antigens can be used as diagnostic and prognostic markers of COVID-19. However, the popular point-of-care biomarker detection technologies, such as the lateral-flow test strips, provide only yes/no information and have very limited sensitivities. Thus, it has a high false negative rate and cannot be used for the quantitative evaluation of patient’s immune response. Conventional ELISA (enzyme-linked immunosorbent assay), on the other hand, can provide quantitative, accurate, and sensitive results, but it involves complicated and expensive instruments and long assay time. In addition, samples need to be sent to centralized labs, which significantly increases the turn-around time. Here, we present a microfluidic ELISA technology for rapid (15-20 minutes), quantitative, sensitive detection of SARS-CoV-2 biomarkers using SARS-CoV-2 specific IgG and viral antigen – S protein in serum. We also characterized various humanized monoclonal IgG, and identified a candidate with a high binding affinity towards SARS-CoV-2 S1 protein that can serve as the calibration standard of anti-SARS-CoV-2 S1 IgG in serological analyses. Furthermore, we demonstrated that our microfluidic ELISA platform can be used for rapid affinity evaluation of monoclonal anti-S1 antibodies. The microfluidic ELISA device is highly portable and requires less than 10 μL of samples for each channel. Therefore, our technology will greatly facilitate rapid and quantitative analysis of COVID-19 patients and vaccine recipients at point-of-care.

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  1. ScreenIT

    SciScore for 10.1101/2020.04.20.052233: (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

    Antibodies
    SentencesResources
    The anti-polyhistidine antibody that was used in polyhistidine-mediated S1 protein immobilization (see Figure S1) was purchased from Thermo Fisher (MA1-21315).
    anti-polyhistidine
    suggested: None
    The horseradish peroxide (HRP) conjugated secondary antibody used in the IgG detection experiment was from the detection antibody in Thermo Fisher’s human total IgG ELISA kit (88-50550-22).
    total IgG
    suggested: (Thermo Fisher Scientific Cat# 88-50550-22, RRID:AB_2574891)
    The HRP conjugation of CR3022, D001, D003, and D006 antibodies were carried out with Abcam’s HRP conjugation kit (ab102890) with a molar ratio of antibody : HRP = 1 : 4.
    D001
    suggested: (Sino Biological Cat# 40150-D001-H, RRID:AB_2857930)
    D006
    suggested: (GenWay Biotech Inc. Cat# GWB-64D006, RRID:AB_10270356)
    In the first step of the reactor preparation process (Figure S2(A)), the working solution of the capture antibody (i.e., D001 in S1 detection experiments) or the anti-His antibody (in IgG detection and antibody affinity experiments) were prepared by diluting the stock solution with the ELISA coating buffer (1× PBS, pH = 7.4) to achieve final concentrations of 10 μg/mL.
    anti-His
    suggested: None
    For the anti-S1 IgG detection experiments (see Figure S2(B) for the detailed protocol), various concentrations of monoclonal antibodies were prepared by diluting the stock solutions with 50 times diluted human serum (the serum was diluted with 1× reagent diluent, which correlates to 1% BSA).
    anti-S1 IgG
    suggested: (Imported from the IEDB Cat# 2E10, RRID:AB_2848047)
    Software and Algorithms
    SentencesResources
    The horseradish peroxide (HRP) conjugated secondary antibody used in the IgG detection experiment was from the detection antibody in Thermo Fisher’s human total IgG ELISA kit (88-50550-22).
    Thermo Fisher’s
    suggested: None

    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 10.1101/2020.04.20.052233v1 on bioRxiv