This volume provides protocols to successfully apply cutting-edge technologies to characterize the biology of T cells at an unprecedented level of complexity. Chapters guide readers through flow cytometry and fluorescence-activated cell sorting, the behaviour of single T cells after adoptive cell transfer (ACT), single cell gene expression by multiplex PCR, lentiviral transduction approaches, protocols to derive large numbers of early-differentiated memory T cells by using dedicated cytokines cocktails, approaches to measure telomerase activity in terminally differentiated T cells, and approaches to define Treg cells at the phenotypic and functional level. The final part of the book is dedicated to the analysis of the differentiation and effector functions of innate T cells, namely the well-known γ/δ T cells, and the recently identified CD8+ mucosal associated invariant T (MAIT) cells. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to theirrespective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.   Authoritative and cutting-edge, T-Cell Differentiation: Methods and Protocols aims to provide protocols that are fundamental to monitor the T cell compartment at the level of single cells in pathological and immunotherapy conditions.