Detection of SARS-CoV-2 epitope-specific CD8⁺ T cell responses in convalescent COVID individuals using HLA-A*02...

As new SARS-CoV-2 subvariants emerge alongside continuous reports of post-acute long-term sequelae after COVID-19, there is an urgent need to mitigate the harmful effects associated with SARS-CoV-2 infection. While antiviral protection is commonly assessed by measuring neutralizing antibodies, specific antiviral CD8⁺ T cell responses are also relevant for long-term immunity. In this context, tools that improve our knowledge regarding anti-SARS-CoV-2 CD8⁺ T cell immunity are important for monitoring antiviral responses, supporting the development of new approaches to induce long-term immunity. Here, we sought to establish a cost-effective platform that enables the direct identification of SARS-CoV-2 peptide-reactive CD8⁺ T cells restricted to HLA-A*02, a highly frequent allele family in the Brazilian population. For this, we produced an HLA-A*02 dimeric molecule (HLA-A*02:01-IgG) in-house, coupled with peptides derived from SARS-CoV-2, and used in flow cytometry assays to identify antigen-specific CD8⁺ T cell populations. Using low concentrations of HLA-A*02:IgG-peptide complex (0.25 μg/mL), we were able to detect anti-SARS-CoV-2-specific CD8⁺ T cells in HLA-A*02-positive individuals, but not in HLA-A*02-negative individuals, as expected. Furthermore, ROC curve analysis showed maximum sensitivity and specificity (100%, AUC = 1.0), highlighting the effectiveness of our method. Overall, our assay identified virus-specific CD8⁺ T cell populations in HLA-A*02 individuals using low concentrations of HLA-A*02:IgG-peptide complex, supporting its use to evaluate the CD8⁺ T cell responses restricted to SARS-CoV-2. This method may provide insights into cellular immunity in the context of the current vaccination and help guide new vaccine and/or therapeutic platform development, including the analysis of memory CD8⁺ T cell subsets.