Diffusion tensor imaging (DTI) is widely used to non-invasively study neural tissue micro-structure. While DTI tractography of large nerve fibers is well accepted, visualization of smaller fibers and resolution of branching fibers is challenging. Sensitivity of DTI to diffusion anisotropy can be further enhanced using long diffusion time that can provide a more accurate representation of the tissue micro-structure. We previously reported that ex vivo fixed brain DTI at long tdiff (192 ms) showed improved sensitivity to fiber tracking compared to short tdiff (48 ms) in 4% formalin-fixed non-human primate (NHP) brains. This study further tested the hypothesis that DTI at longer diffusion time improves DTI fiber tracking in the in vivo NHP brains on a clinical 3 Tesla MRI scanner. Compared to fixed brains, the in vivo ADC was larger by a factor of 5. Also, the white-matter FA was 28% higher in the in vivo study as compared to our ex vivo experiments. Compared to short tdiff, long tdiff increased white-matter FA by 6.0±0.5%, diffusion was more anisotropic, tensor orientations along major fiber tracts were more coherent, and tracked fibers were about 10.1±2.9% longer in the corpus callosum and 7.3±2.8% longer along the cortico-spinal tract. The overall improvements in tractography were, however, less pronounced in the in vivo brain than in fixed brains. Nonetheless, these in vivo findings reinforce that DTI tractography at long diffusion time improves tracking of smaller fibers in regions of low fractional anisotropy.