TY - JOUR
T1 - Development of an in-house SARS-CoV-2 interferon-gamma ELISpot and plate reader-free spot detection method
AU - Mak, Willem A
AU - Koeleman, Johannes G M
AU - Ong, David S Y
N1 - Funding Information:
This research was funded by the Franciscus Gasthuis & Vlietland Hospital, Rotterdam, the Netherlands and did not receive any additional funding from agencies in the public, commercial, or not-for-profit sectors.
Publisher Copyright:
© 2021 Elsevier B.V.
PY - 2022/2
Y1 - 2022/2
N2 - Coronavirus disease 2019 (COVID-19) vaccination programs rolled out in an attempt to stop the COVID-19 pandemic. Besides neutralising antibodies, effective T cell responses are also crucial for protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and COVID-19 disease severity. To assess SARS-CoV-2-specific T cell immunity, we developed an interferon-gamma (IFN-γ) enzyme-linked immunospot (ELISpot) that can be deployed in research and diagnostic settings. We optimised our ELISpot by testing multiple antigen concentrations to stimulate peripheral blood mononuclear cells of SARS-CoV-2-unexposed, COVID-19 convalescent and COVID-19 vaccinated volunteers. Also, we developed an ELISpot plate reader-free method to detect and quantify spots, which we compared to manual spot counting and automated analysis by an ELISpot plate reader. We observed strong SARS-CoV-2-reactive T cell responses in COVID-19 convalescent, and COVID-19 vaccinated volunteers but absent or only weak responses in unexposed volunteers. Overall, antigens with concentrations from 0.1 to 5.0 μg/mL per peptide elicited similar T cell responses. Also, our plate reader-free detection method reliably detected and quantified SARS-CoV-2-specific T cells, demonstrated by an excellent reliability when compared to manual analysis and automated analysis by an ELISpot plate reader.
AB - Coronavirus disease 2019 (COVID-19) vaccination programs rolled out in an attempt to stop the COVID-19 pandemic. Besides neutralising antibodies, effective T cell responses are also crucial for protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and COVID-19 disease severity. To assess SARS-CoV-2-specific T cell immunity, we developed an interferon-gamma (IFN-γ) enzyme-linked immunospot (ELISpot) that can be deployed in research and diagnostic settings. We optimised our ELISpot by testing multiple antigen concentrations to stimulate peripheral blood mononuclear cells of SARS-CoV-2-unexposed, COVID-19 convalescent and COVID-19 vaccinated volunteers. Also, we developed an ELISpot plate reader-free method to detect and quantify spots, which we compared to manual spot counting and automated analysis by an ELISpot plate reader. We observed strong SARS-CoV-2-reactive T cell responses in COVID-19 convalescent, and COVID-19 vaccinated volunteers but absent or only weak responses in unexposed volunteers. Overall, antigens with concentrations from 0.1 to 5.0 μg/mL per peptide elicited similar T cell responses. Also, our plate reader-free detection method reliably detected and quantified SARS-CoV-2-specific T cells, demonstrated by an excellent reliability when compared to manual analysis and automated analysis by an ELISpot plate reader.
KW - COVID-19
KW - Diagnostic
KW - ELISpot
KW - SARS-CoV-2
KW - T cells
UR - http://www.scopus.com/inward/record.url?scp=85120577558&partnerID=8YFLogxK
U2 - 10.1016/j.jviromet.2021.114398
DO - 10.1016/j.jviromet.2021.114398
M3 - Article
C2 - 34863782
SN - 0166-0934
VL - 300
JO - Journal of Virological Methods
JF - Journal of Virological Methods
M1 - 114398
ER -