The regulation of T cell activation and immune synapse formation is poorly understood at the molecular level. CD45, a receptor-like protein tyrosine phosphatase (RPTP), is a key regulator of T cell activation. CD45 activity may be controlled through changes in: receptor localization on the cell surface; receptor associations with other membrane proteins, membrane lipids (e.g., microdomains), and intracellular signaling pathways; and receptor lateral mobility. Using single particle tracking, we demonstrate that activation of Jurkat T cells significantly decreases CD45 lateral mobility. Dominant peptide inhibition experiments show that CD45 binding to spectrin-ankyrin is partially responsible for CD45 immobilization.  Other potential mechanisms of CD45 immobilization remain to be identified.