An efficient immunoassay for the B cell help function of SARS-CoV-2-specific memory CD4+ T cells

  1. Asgar Ansari
  2. Shilpa Sachan
  3. Bimal Prasad Jit
  4. Ashok Sharma
  5. Poonam Coshic
  6. Alessandro Sette
  7. Daniela Weiskopf
  8. Nimesh Gupta

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

No abstract available

Article activity feed

  1. Version published to 10.1016/j.crmeth.2022.100224
  2. ScreenIT

    SciScore for 10.1101/2021.08.12.21261970: (What is this?)

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

    Table 1: Rigor

    EthicsIRB: Human Blood Sample: The study was approved by the Institutional Review Boards of National Institute of Immunology (NII) and All India Institute of Medical Sciences (
    Consent: Informed consent was obtained from all the donors/subjects during the enrolment.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    For all the autologous assays, PBMCs after revival were surface stained with antibody cocktail for 15-20 minutes at 4ºC in dark: fixable viability dye efluor 506, anti-CD4 AF700 (RPA-T4)
    anti-CD4
    suggested: (SouthernBiotech Cat# 1540-27, RRID:AB_2794844)
    Soluble growth factors/cytokines in B+M co-cultures were blocked using 10μg/mL of following neutralizing antibodies: purified anti-BAFF (#AF124) and anti-SCGF-β (#MAB1904) (R&D Systems)
    anti-BAFF
    suggested: None
    anti-SCGF-β
    suggested: None
    After incubation, cells were stained with the following antibody cocktails: fixable viability dye efluor 506, anti-CD3 BV650 (UCHT1)
    anti-CD3
    suggested: None
    UCHT1
    suggested: None
    For calculating the concentration of SARS-CoV-2 spike specific IgG antibody, purified SARS-CoV-2 neutralizing antibody (CR3022) was used for plotting the standard.
    SARS-CoV-2 spike specific IgG
    suggested: None
    CR3022
    suggested: None
    For detection of ASC spots, anti-human IgG-550 (MT78/145) and IgM-640 (MT22) detection antibodies (Mabtech) were added and incubated for 2 hours at RT in dark.
    anti-human IgG-550
    suggested: None
    IgM-640 ( MT22
    suggested: None
    Software and Algorithms
    SentencesResources
    Data were analyzed using FlowJo 10.3.0.
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Statistical analyses and data visualization were performed with the GraphPad Prism software version v8.4.0.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)

    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:
    Certainly, monocyte supplemented T-B co-cultures overcome these limitations by consistently enhancing the plasma cell output and antibody production in co-cultures stimulated with CoV-S protein and peptides. Moreover, the improved sensitivity in the B-cell output in monocyte supplemented settings was accompanied by the enhanced magnitude of functional activated CD4+ T cells, which was associated with enhanced level of TCR directed effector cytokines like IL-2, IFN-γ, IL-4, IL-10, IL-17 and sCD40L (Geginat et al., 2001; Rochman et al., 2009). Monocytes also provided crucial cytokines like IL-15 that seems to promote the survival and proliferation of T cells in these spike-specific co-cultures (Dooms et al., 1998; Geginat et al., 2001). These observations also highlighted the fact that these COVID-19 recovered subjects harbor spike-specific memory CD4+ T cells that are capable of inducing B cell differentiation into ASCs and produce IgG during any recall response. Monocytes and B cells act as antigen presenting cells to prime and activate the memory CD4+ T cells (Hong et al., 2018; Hua and Hou, 2020; Randolph et al., 2008; Tacke et al., 2006). However, they differ in their abilities to deliver the secondary activation signals as evident by the constitutive expression of CD86 on all classical monocytes, but not on the B cells. In fact, signals from activated T cells like IL21 is essential for inducing the co-stimulatory ligands on the B cells (Attridge et al., 2014). Besides, mo...

    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.

    Results from scite Reference Check: We found no unreliable references.

    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.

  3. Version published to 10.1101/2021.08.12.21261970v1 on medRxiv