High temporal resolution path tracking highlights intratumoral heterogeneity of speed and directionality of tumor cell migration in focal micro-regions. (A) 10x field view of tumor cells in GBM-8 labeled by transduction of an MMLV-retroviral vector constitutively expressing ZsGreen, and (B) accompanying migration paths after 11 hours of imaging. Scale bars represent 50 μm. Tumor cell location was tracked every 11 minutes. Cells with long, direct paths are observed migrating past stationary cells. (C) Cell speeds are distributed log-normally (R2 = 0.95) in this representative population (GBM-8). The red line represents the best fit regression to a log-normal base model. (D) Correlation between directionality and speed in a representative tumor (GBM-8, r = 0.65, p < 0.0001). Similar results were obtained in 5 out of 7 tumors analyzed (Spearman r correlation coefficient ranged from 0.45 to 0.65). (E) A representative tumor cell tracked over 10 hours, in 55-minute intervals, demonstrating rapid cell division during migration (GBM-13), coupled with the instantaneous speed over migration time for this cell (right).