T Cell ID: Identification and quantification of T cell responses
Immunitrack offers a powerful platform for finding, validating and quantifying T cell responses towards cancer neo-epitopes and viral derived epitopes.
Step 1. Selection of potential T cell epitopes based on partner data or literature. Alternatively, and in most cases, we select candidate epitopes based on our NeoScreen Platform, in which we can screen hundreds of epitopes on multiple HLA Class I and Class II alleles.
Step 2: Confirm and validate epitopes after long term stimulation of PBMCs followed by flow cytometry analysis of effector and activation markers on T cells.
Step 3. Quantification of specific T cells by Tetramer stain.
In this application note you will find a platform developed by Immunitrack to identify and quantify specific T cell responses from PBMC samples.
The expression of the highly polymorphic T cell receptor (TCR) allows T cells to specifically recognize foreign and self-antigens presented in the context of Major Histocompatibility Complex (MHC). The tremendous diversity of the TCR repertoire (up to 1020) together with the high diversity of matching MHC molecules in humans (also called HLA – human leukocyte antigen), is the hallmark of the adaptive immune system but simultaneously poses technical challenges for identification of specific T cell responses in the context of cancer, viral infections, or autoimmunity.
To tackle this, we have developed a methodology (NeoScreen) to narrow down the list of potential immunogenic peptides (epitopes) based on the stability of the peptide-MHC complex. To validate epitopes, we use PBMCs from healthy volunteers or diseased patients, which we stimulate with peptide (or peptide pools) and then use flow cytometry as readout. We can stain for the numerous activation markers that are expressed on the surface of specific T cells. Moreover, we can perform intracellular cytokine stain (ICS) for detection of effector cytokines.
To identify and quantify specific T cells we can synthesize a library of MHC-peptide tetramers, since we have a large collection of HLA-I and HLA-II molecules in house. Tetramers are fluorescently labeled, and samples are stained for flow cytometry analysis and/or T cell sorting.
As an example, we present a case study in which we identified Spike-specific CD8 T cells in SARS-Cov-2 vaccinated and convalescent donors.