AbstractMany biological materials are hierarchically structured, with highly anisotropic structures and properties on several length scales. To characterize the mechanical properties of such materials, detailed testing methods are required that allow precise and site-specific measurements on several length scales. We propose a fracture toughness measurement technique based on notched focused ion beam prepared cantilevers of lower and medium micron size scales. Using this approach, site-specific fracture toughness values in dental enamel were determined. The usefulness and challenges of the method are discussed.