Early immune response in mice immunized with a semi-split inactivated vaccine against SARS-CoV-2 containing S protein-free particles and subunit S protein

  1. Marek Petráš
  2. Petr Lesný
  3. Jan Musil
  4. Radomíra Limberková
  5. Alžběta Pátíková
  6. Milan Jirsa
  7. Daniel Krsek
  8. Pavel Březovský
  9. Abhishek Koladiya
  10. Šárka Vaníková
  11. Barbora Macková
  12. Dagmar Jírová
  13. Matyáš Krijt
  14. Ivana Králová Lesná
  15. Věra Adámková

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Abstract

The development of a vaccine against COVID-19 is a hot topic for many research laboratories all over the world. Our aim was to design a semi-split inactivated vaccine offering a wide range of multi-epitope determinants important for the immune system including not only the spike (S) protein but also the envelope, membrane and nucleocapsid proteins. We designed a semi-split vaccine prototype consisting of S protein-depleted viral particles and free S protein. Next, we investigated its immunogenic potential in BALB/c mice. The animals were immunized intradermally or intramuscularly with the dose adjusted with buffer or addition of aluminum hydroxide, respectively. The antibody response was evaluated by plasma analysis at 7 days after the first or second dose. The immune cell response was studied by flow cytometry analysis of splenocytes. The data showed a very early onset of both S protein-specific antibodies and virus-neutralizing antibodies at 90% inhibition regardless of the route of vaccine administration. However, significantly higher levels of neutralizing antibodies were detected in the intradermally (geometric mean titer - GMT of 7.8 ± 1.4) than in the intramuscularly immunized mice (GMT of 6.2 ± 1.5). In accordance with this, stimulation of cellular immunity by the semi-split vaccine was suggested by elevated levels of B and T lymphocyte subpopulations in the murine spleens. These responses were more predominant in the intradermally immunized mice compared with the intramuscular route of administration. The upward trend in the levels of plasmablasts, memory B cells, Th1 and Th2 lymphocytes, including follicular helper T cells, was confirmed even in mice receiving the vaccine intradermally at a dose of 0.5 μg.

We demonstrated that the semi-split vaccine is capable of eliciting both humoral and cellular immunity early after vaccination. Our prototype thus represents a promising step toward the development of an efficient anti-COVID-19 vaccine for human use.

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  1. ScreenIT

    SciScore for 10.1101/2020.11.03.366641: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIACUC: All protocols were approved by the Animal Care Committee of the Institute for Clinical and Experimental Medicine and the Ministry of Health of the Czech Republic.
    Randomizationnot detected.
    Blindingnot detected.
    Power AnalysisA >80% power of the test was only achieved when comparing the geometric mean titers of antibodies between immunized and unimmunized mice.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    After incubation, the wells were washed and a horseradish peroxidase-conjugated anti-S antibody added, producing a sandwich complex.
    anti-S
    suggested: None
    ELISA antibodies assay: Specific anti-SARS-CoV-2 IgG antibodies in mice were detected with a commercial ELISA kit (Human Anti-2019 nCoV(S) IgG ELISA Kit, FineTest, Wuhan, China) where the purified horseradish peroxidase (HRP)-conjugated anti-mouse IgG detection antibodies were replaced by HRP-conjugated AffiniPure goat anti-mouse IgG (Wuhan Fine Biotech Co.
    anti-SARS-CoV-2 IgG
    suggested: None
    Anti-2019
    suggested: None
    , Ltd. China) diluted with an antibody dilution buffer at 1:1,000 according to the anti-mouse IgG manufacturer’s instructions.
    anti-mouse IgG
    suggested: None
    Statistical analysis: Anti-S specific IgG antibodies as well as virus-neutralizing antibodies were expressed as the titer.
    Anti-S specific IgG
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Serial 2-fold dilutions of the plasma from 1:2 to 1:32 were subsequently incubated with 100 TCID50 of the vaccine of the SARS-CoV-2 virus strain for 2 hours and added to VERO E6 cell monolayers.
    VERO E6
    suggested: RRID:CVCL_XD71)
    Experimental Models: Organisms/Strains
    SentencesResources
    A total of 52 BALB/c mice (Velaz, Czech Republic) were immunized with vaccine (40 mice) or placebo (12 mice) at age 9-10 weeks.
    BALB/c
    suggested: None
    Software and Algorithms
    SentencesResources
    Statistical analyses were performed using Prism 8 (GraphPad Software, Inc.
    GraphPad
    suggested: (GraphPad Prism, RRID:SCR_002798)
    San Diego, CA) and STATA version 16 software (StatCorp, College Station, TX).
    STATA
    suggested: (Stata, RRID:SCR_012763)

    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 10.1101/2020.11.03.366641 on bioRxiv