The intricate connectivity and molecular identity of axons is maintained for decades, but also adapts to changes in neuronal activity and environment.The axon fulfills these paradoxical demands thanks to a compartmented cytoskeletal architecture that ensures the transport, anchoring and mobility of axonal components. We can now directly visualize these molecular assemblies in situ, thanks to the development of optical super-resolution microscopy techniques (Leterrier et al., 2017). We have used STochastic Optical Reconstruction Microscopy (STORM) to map the nano-architecture of cytoskeletal and scaffold assemblies within the axon. In the axon initial segment, a key compartment for the maintenance of neuronal polarity, we quantitatively resolved a highly organized actin/spectrin/ankyrin scaffold (Leterrier et al., 2015). In addition, we localized several new components within this scaffold (Huang et al., 2017; Berger et al., 2018). Finally, we are developing new versatile labeling, acquisition and processing strategies to characterize the nanoscale architecture of the axon, bringing insights on how it organizes axonal transport (Culley et al., 2018; Almada et al., 2018).