Production of Antigen-Specific Human CD4⁺ T Cell Lines and Clones

Methodologies to generate single antigen-specific T cells are based on the T cell specificity, activation, or other subsequent functional measures. One of the most powerful tools to isolate human CD4⁺ T cell clones is utilization of MHC Class II tetramers. Flow cytometer-based tetramer technology mimics the recognition of the specific antigenic peptide in the context of HLA class II (tetramer) by the T cell receptor. MHC class II tetramers, which can be exogenously loaded to contain any peptide of interest that binds to them (T cell epitopes), provide a valuable tool for detection of T cells in the peripheral blood or the tissue that are specific for antigens from different viruses, tumors, or self-proteins (autoimmunity). Generation of T cell clones with a defined antigen specificity allows for a deeper characterization and functional assessment at single cell level. This is important for determination of the epitope specificity and functional phenotype of the disease associated T cells. Single cell cloning can be utilized in the direct sequencing of the T cell receptor alpha/beta pairs that are prevalent in the disease and therefore provides a platform for T cell receptor engineering, which has applications in the immunotherapy.