The Leica EM AFS2 freeze substitution device is used for low temperature embedding of samples prepared by HPF. The process of freeze substitution involves three phases. At first, the water of a vitrified specimen is exchanged against an organic solvent like acetone or ethanol. During this step, the structure is additionally fixated by staining agents. Afterwards, the organic solvent is then exchanged by a resin followed by curing the resin either thermally or by UV irradiation. The EM AFS2 freeze substitution device comes with an automatic reagent handling system (EM FSP) which facillitates long FS protocols.