Nanoscopic biomechanical properties: indentation modulus, hardness and damage behaviorThe region-dependent chemical composition and microstructural organization of the spike cuticle are reflected into spatially varying mechanical properties, as measured by depth sensing nanoindentation (Figure 5 A). The outer FAP-dominated region is clearly the stiffest and hardest location (indentation modulus of 64.9 ± 3.5 GPa and hardness of 4.0 ± 1.0 GPa). Both mechanical parameters show a considerable drop (almost a factor of 2 for the modulus and of 5 for hardness) in the thin outer helicoidal region, which displays the lowest values of the entire spike, consistent with the low mineral content and compositional modifications. Moving to the striated region, indentation modulus and hardness increase and the former also shows slightly higher values with respect to the inner twisted plywood (Table 1), which is conceivable considering the highly ordered arrangement of the fibers. Furthermore, a small negative gradient of about 4 GPa is observed in the indentation modulus in this location, probably reflecting the corresponding decrease in Ca content. To better characterize the spatial tuning in local mechanical behavior from the heavily mineralized to the parallel fibers region, we performed high resolution nanoindentation with a sharp cube corner probe (Figures 5B-D and Figure S8 ). The high-resolution maps confirm the unusual drop in modulus within the outer twisted plywood region, with values slightly below 20 GPa and no differences between longitudinal and transverse sections, consistent with the isotropic nature of the twisted plywood organization. In particular, the transition region (featuring an individual lamella with fairly high mineral content) has a clear signature in the mechanical maps (Figures 5C and D) with a modulus of about 40-45 GPa, allowing a gradual switch from the highly mineralized to the outer helicoidal region. One additional feature of the highly mineralized region is a quite heterogeneous mechanical behavior (Figure S9 ), likely reflecting the spatial variations of phosphate intensity and crystallinity measured by Raman spectroscopy. A similar mechanical modulation featuring stiffer crystalline domains and less stiff amorphous islands has been reported for the heavily mineralized impact region of the spearer appendage [35]. Although a thin outer helicoidal region enclosing highly co-aligned fibers has been found in other biological penetrating tools such as the uropod back spike of the mantis shrimp [42] and the spider fang [45], a novel feature observed here is that this region, sandwiched between the hard outer cover and the stiff parallel-fiber region, presents substantially lower mechanical properties (modulus and hardness) than the neighborhood.