Molecular diffusion plays an important role in many biological phenomena. Magnetic Resonance (MR) imaging is inherently sensitive to diffusion and can be used to help understand diffusion processes. Diffusion MR imaging is most widely used for imaging the ischemic brain. Diffusion imaging and diffusion tensor imaging (DTI) have also found clinical application in areas such as tumor characterization throughout the body, imaging of demyelinating disorders, and fiber tract mapping. DTI is also now widely used in small animal imaging-both in vivo and in characterizing excised tissue. DTI studies in these settings can be accomplished with high resolution and can offer exquisite contrast, but the technical and practical challenges can sometimes be different than those seen on clinical MRI scanners. Here, a stepwise methodology is presented for using small-bore, high field strength scanners (>3 T) for DTI. This chapter is aimed at addressing readers with no prior knowledge of DTI and we present both a basic explanation of underlying principles and a practical approach to the experiment.