Measles viruses have shown potent oncolytic activity as a therapeutic against a variety of human cancers in animal models and are currently being tested in clinical trials in patients. In contrast to using measles virus as a vaccine, oncolytic activity depends on high concentrations of infectious virus. For use in humans, the high-titer measles virus preparations must also be purified to remove significant levels of cellular proteins and nucleic acid resulting from the cytolytic products of measles virus replication and release. Pleomorphic measles virus must be treated as >1-μm particles that are extremely shear sensitive to maximize recoveries and retain infectivity. Therefore, to maximize the recovery of sterile, high titer infectious measles viruses, the entire production and purification process must be done using gentle conditions and aseptic processing. Here we describe a procedure applicable to the production of small (a few liters) to large (50–60 L) batches of measles virus amplified in Vero cells adapted to serum-free growth. Cell culture supernatant containing the measles virus is clarified by filtration to remove intact Vero cells and other debris, and then treated with Benzonase® in the presence of magnesium chloride to digest contaminating nucleic acid. The measles virus in the treated cell culture supernatant is then concentrated and purified using tangential flow filtration (TFF) and diafiltration. The concentrated and diafiltered measles virus is passed through a final clarifying filter prior to final vialing and storage at <−65°C. An infectivity assay to quantify infectious measles virus concentration based on the TCID50 method is also described. This procedure can be readily adapted to the production and purification of measles viruses using good manufacturing practices (GMP).