Transmissible spongiform encephalopathies (TSE) or prion diseases exhibit strain variation, a phenomenon that has been studied extensively in mouse bioassays. Despite the introduction of many rapid in vitro systems, bioassays remain a key tool in defining prion strains and their ability to transmit disease in vivo. Prion strains can be characterized by a range of phenotypic characteristics such as incubation period, vacuolar pathology, and distribution of the abnormal form of PrP following experimental transmission of the agent into a panel of mice (transgenic or wild type). Interpretation of these characteristics requires considerable experience and an understanding of the procedures used to define them. This chapter reviews the techniques used in strain typing of prion diseases from inoculum preparation and pathological studies to data interpretation alongside an extensive troubleshooting guide.