Over the past decade, diffusion tensor imaging (DTI) has offered researchers and clinicians a new noninvasive window into the developing human brain, from preterm infants through adolescents and young adults. DTI improves on conventional MR imaging, such as T1-weighted and T2-weighted sequences, through its sensitivity to many microstructural features of neural organization. This has enabled visualization of the early cerebral laminar architecture in premature infants, of developing white matter before myelination, and of the microarchitecture of the cerebral cortex during preterm maturation. DTI provides reproducible quantitative measures, such as mean diffusivity and fractional anisotropy, that reflect the underlying tissue properties of gray matter and white matter and may therefore become useful as developmental milestones for the improved assessment of abnormal brain maturation. Furthermore, three-dimensional fiber tractography based on DTI can reveal the developing axonal connectivity of the human brain as well as aberrant connectivity in structural brain malformations. In this article, applications of DTI and fiber tractography to the study of human brain development are reviewed. The new insights into brain maturation afforded by DTI promise to improve the diagnostic evaluation of an array of congenital, metabolic, and neurodevelopmental disorders.