A study by Sandberg and colleagues investigated several immune parameters during the acute phase and two follow-ups 5 and 9 months after COVID-19 disease in hospitalized patients. To investigate the development and persistence of virus-specific cellular and humoral immune responses, Mabtech’s FluoroSpot assays were chosen.
Memory B cells were analyzed by combining pre-stimulation with antigen-specific IgA and IgG detection in FluoroSpot. For this, recombinantly produced SARS-CoV-2 S1 or N proteins were coated onto a FluoroSpot plate before adding the previously activated cells. Compared with pre-pandemic samples, almost every tested patient sample included S1- or N-specific memory B cells. Interestingly, these antigen-specific cells in relation to the total number of IgG secreting cells had very variable ratios across patients.
In order to study antigen-specific T cell responses, the authors chose FluoroSpot Plus: Human IFN-γ/IL-2/TNF-α, acknowledging that polyfunctional T cells contribute to a robust antiviral response (T cells are considered polyfunctional for example when they secrete several cytokines simultaneously). Every patient in the cohort developed a polyfunctional T cell response to at least one of the SARS-CoV-2 derived peptide pools tested. For the single-secreting responses, a clear difference between healthy controls and patients was seen for IFN-γ and IL-2. To assess an additional level of information, a measurement of the relative amounts of secreted cytokines, the relative spot volume (RSV) feature of Mabtech IRIS, was used with the aim to further dissect the polyfunctional T cell response. In this study, cells secreting more than one type of cytokine also secreted a higher relative amount of each cytokine.
This study is an example of how polyfunctional T cells can be studied with FluoroSpot and how the relative spot volume measurement can be included in the analysis. One could speculate whether the reason behind polyfunctional T cells being important in antiviral defense stems from secreting several cytokines or larger amounts of cytokines or simply a combination thereof. We are looking forward to more studies published where researchers incorporate RSV in their analysis of FluoroSpot experiments.
