Expansion microscopy (ExM) is a recently developed optical superresolution approach based on the original idea that physical separation of light point sources will overcome the limits of the conventional-diffracted microscope. Indeed, an expansion factor of 4 will give for example a lateral resolution of around 70nm compared to the 270 nm resolution of the non-expanded sample. The tissue samples (cerebellum) will originate from mice injected in the spinal cord with an AAVvirus that express tdTomato. The targeted neurons project then to the cerebellum, allowing detection of specific synapses (mossy fibers) in that brain region. Our goal will be to observe the fine structure of the synapses and its internal organization. We will demonstrate the gain of resolution obtained using ExM. No special pre-requisite is needed to attend this workshop. In order to facilitate the starting of such a project we will go through all the possible difficulties a beginner could encounter. In this workshop we will present a step by step detailed protocol based on the proExM (protein retention ExM) method published by Asano et al. The specimen (here a tissue slice) in which proteins have been covalently anchored with acryloyl-X, SE (AcX), is embedded in an acrylamide/bis-acrylamide/sodium acrylate gel (hydrogel). Then, a homogenization /disruption step is performed by enzymatic digestion or by a high temperature treatment in presence of detergent. Finally, using the swellable property of sodium acrylate, the gel is expanded with water undergoing an isotropic three-dimensional expansion by 4-4.5 times in that case. To go further we will also discuss about recent developments allowing even a better resolution with expansion factors of 10 or more.