Cytotoxic T lymphocytes (CTLs) are important constituents of the adaptive immune system. Development of CTLs are particularly important for bacterial and viral infections, in addition to tumor surveillance. Measuring T cell immune function is important in evaluating host defense, allergy, autoimmunity, transplant rejection, and tumor immunity. In these recent years it has become possible to measure multiple effector functions in a single cell such as cytokine, transcription factors, and cytolytic function. In addition these parameters can be evaluated in conjunction with cellular proliferation. In this chapter we detail these cellular based assays and the methods used to characterize and quantify both phenotype and function of CTL.