Background Evaluating the antigen-specific cellular immune response in multiple cellular compartments is important for understanding anti-tumor immunity and development of personalized vaccines and immunotherapy.

Investigation of other T cell types than the classically antigen-specific CD8 and CD4 T cells, such as iNKT and MAIT have increased interest in cancer.

Analysis of single cell surface proteins, transcriptome and TCR/BCR gene clonotypes, enables understanding of the antigen-specific recognition at the immune synapse which is key in understanding the antigen- specific cellular immune response in infectious diseases as well as cancers.

Methods The dCODE Dextramer® technology together with BD Rhapsody™ Single-Cell Analysis combines genomic profiling with antigen-specific recognition allowing deep pheno- and genotypic analysis of antigen-specific T and B cells. However, to unveil the heterogenicity of the cellular immune responses seen in cancer and infectious disease many specificities need to be investigated in multiple donors to get the complete picture.

Results Here we report how multiple antigen-specific T- and B-cell responses in multiple samples can be analyzed simultaneously in one run using sample tagging. The mixed sample was stained with a panel of dCODE Dextramer and antibody reagents followed by single cell analysis. The mixed sample, consisted of PBMCs from a previously SARS-CoV-2 infected and vaccinated individual, a donor previously exposed to EBV and CMV, as well as an sample stimulated EBV and Tetanus toxoid peptides.

Evaluation of the conventional T-cell responses consisted of 8 different MHCI, 2 different MHCII, and 4 negative control dCODE Dextramer® (RiO) reagents. SARS-CoV-2 specific B cells were analyzed using dCODE Klickmer® carrying SARS-CoV-2 Spike protein. iNKT cells and MAIT cells were analyzed using CD1d and MR1 dCODE Dextramer reagents.

The BD® AbSeq Immune Discovery Panel identifying 30 immune related surface proteins was employed for immune phenotyping.

Conclusions We demonstrate a method to do deep characterization of the complete antigen-specific cellular immune response in multiple donor samples at scale, by applying combined T- and B-cell antigen-specific detection, alongside Identification of full length VDJ TCR and BCR clonotypes, targeted mRNA expression, and immune cell surface marker phenotyping.

For Research Use Only. Not for use in therapeutic or diagnostic procedures

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