Seizure-related Protein 6 (Sez6) is involved in neuronal development and synaptic function. Sez6 is cleaved near its transmembrane region by the protease BACE1, producing a soluble protein (sSez6). This shed sSez6 is detected in cerebrospinal fluid (CSF) and sSez6 CSF levels are altered in multiple neurological diseases and disorders. We aim to understand the cellular processes that affect sSez6 shedding from neurons, in addition to the functional consequences of this change. In this study, immunocytochemical analysis revealed that endogenous Sez6 is localised in the Golgi, early, late and recycling endosomes and at the cell surface of cultured rat hippocampal neurons. We used the Retention Using Selective Hooks (RUSH) system to synchronise and follow the anterograde trafficking of Sez6 in real time. HeLa cells and hippocampal neurons were transfected with the Sez6 RUSH plasmid. Newly synthesized Sez6-RUSH is retained in the endoplasmic reticulum and is released by the addition of biotin. Using spinning-disk and super-resolution microscopy to image live and fixed cells, Sez6 trafficking was examined for up to 120 minutes after release. Newly synthesized SBP-GFP-Sez6 was first concentrated in the Golgi apparatus before being distributed in vesicles throughout the soma and neuronal dendrites. SBP-GFP-Sez6 cell surface staining tended to increase from 45 to 120 minutes post release. Further work will examine the effect of altered BACE1 activity on Sez6 localisation and trafficking in neuronal subcellar compartments. These results provide insight into the mechanisms of sSez6 production and increase our understanding of molecular factors contributing to altered Sez6 levels in disease.