The three-dimensional positions of immune cells can be tracked in live tissues precisely as a function of time using two-photon microscopy. However, standard methods of analysis used in the field and experimental artifacts can bias interpretations and obscure important aspects of cell migration such as directional migration and non-Brownian walk statistics. Therefore, methods were developed for minimizing drift artifacts, identifying directional and anisotropic (asymmetric) migration, and classifying cell migration statistics. These methods were applied to describe the migration statistics of CD8+ T cells in uninflamed lymph nodes. Contrary to current models, CD8+ T cell statistics are not well described by a straightforward persistent random walk model. Instead, a model in which one population of cells moves via Brownian-like motion and another population follows variable persistent random walks with noise reproduces multiple statistical measures of CD8+ T cell migration in the lymph node in the absence of inflammation. 