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Abstract: Several prevalent and life-threatening agents enter the organism via the mucosa. In this case, a mucosal cellular immune response is essential for protection and is therefore considered the main objective of vaccination. The frequency of antigen-specific CD4+ and CD8+ T cells can be determined directly in human whole blood by a combination of surface marker and intracellular cytokine staining. Immune cells primed in the mucosal compartment also migrate through the blood and can be identified by expression of the gut-specific homing receptor α4β7. Simultaneously, these lymphocytes can be functionally characterized regarding their differentiation status by analysis of CD45RO and CD27 expression and effector functions by measuring intracellular perforin or granzyme B content. Thus, the technique described in the paper is a powerful tool for clinical monitoring of the total cellular immune response to complex antigens during infection or vaccination. [Copyright &y& Elsevier]