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Detection of SARS-CoV-2-Specific Cells Utilizing Whole Proteins and/or Peptides in Human PBMCs Using IFN-ƴ ELISPOT
Madeleine M Rasche1, Ella C Kaufmann1, Tamar Ratishvili1
1Mayo Clinic Vaccine Research Group, Mayo Clinic, Rochester, MN, USA.
Abstract:
SARS-CoV-2 continues to threaten global public health, making COVID-19 immunity studies of utmost importance. Waning of antibody responses postinfection and/or vaccination and the emergence of immune escape variants have been ongoing challenges in mitigating SARS-CoV-2 morbidity and mortality. While a tremendous amount of work has been done to characterize humoral immune responses to SARS-CoV-2 virus and vaccines, cellular immunity, mediated by T cells, is critical for efficient viral control and protection and demonstrates high durability and cross-reactivity to coronavirus variants. Thus, ELISPOT, a standard assay for antigen-specific cellular immune response assessment, allows us to evaluate SARS-CoV-2-specific T-cell response by quantifying the frequency of SARS-CoV-2-specific cytokine-secreting cells in vitro. We have outlined a detailed procedure to study T-cell recall responses to SARS-CoV-2 in human peripheral blood mononuclear cells (PBMCs) following infection and/or vaccination using an optimized IFN-γ ELISPOT assay. Our methodologies can be adapted to assess other cytokines and are a useful tool for studying other viral pathogen and/or peptide-specific T-cell responses.
Insights
Investigating T-cell immunity to SARS-CoV-2 is crucial for understanding COVID-19 protection. This study details an optimized Interferon-gamma ELISPOT assay to measure T-cell responses after infection or vaccination.
Area of Science:
- Immunology
- Virology
- Infectious Diseases
Background:
- SARS-CoV-2 poses an ongoing global health threat, necessitating robust immunity assessments.
- Waning antibody responses and immune escape variants complicate COVID-19 mitigation efforts.
- Cellular immunity, particularly T-cell responses, is vital for durable viral control and cross-variant protection.
Purpose of the Study:
- To outline a detailed protocol for assessing SARS-CoV-2-specific T-cell recall responses.
- To evaluate T-cell immunity in individuals post-infection and/or vaccination.
- To establish a reliable method for studying cellular immune responses to viral pathogens.
Main Methods:
- Utilized an optimized Interferon-gamma (IFN-γ) ELISPOT assay.
- Employed human peripheral blood mononuclear cells (PBMCs).
- Quantified SARS-CoV-2-specific cytokine-secreting T cells in vitro.
Main Results:
- Established a detailed procedure for T-cell response assessment.
- Demonstrated the utility of the IFN-γ ELISPOT assay for SARS-CoV-2 immunity studies.
- Highlighted the adaptability of the methodology for other cytokines and pathogens.
Conclusions:
- The optimized IFN-γ ELISPOT assay provides a valuable tool for evaluating SARS-CoV-2-specific T-cell immunity.
- This methodology supports the study of cellular immune memory durability and breadth.
- The protocol can be adapted for broader applications in infectious disease research.
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