The generation of a specific immune response and an immunological memory which confer protective immunity against a wide range of various pathogens and tumors is based on the immune system´s ability to form and maintain antigen specific B- and T-lymphocytes. These antigen specific lymphocytes specifically recognize pathogenic and tumorigenic structures through their unique antigenic receptors (so called B- and T-cell receptors) thereby limiting their effector functions to a single pathogenic or tumorigenic structure (epitope). In order to cope with the enormous number of diverse potential pathogens the immune system has to generate a vast number of T- and B-lymphocytes with their unique T- and B-cell receptors. The detection and characterization of these rare antigen specific T- and B cells is therefore essential for the elucidation of the status of the protective immunity and the factors which contribute to the immune system´s capability to control pathogen and tumor growth. For the detection and analysis of ex vivo antigen-specific CD8+ cytotoxic T cells we are using the MHC class I-peptide Streptamer technology. In addition, functional characterization of antigen-specific CD4+ T-helper cells and CD8+ cytotoxic T cells is performed by intracellular cytokine staining (ICS) following provocation with antigen of interest. Both analyses are performed using multi-parametric flow cytometry which allows characterization of T cell subsets in great detail.