The plant endomembrane system controls transport of diverse cargo including proteins, lipids, carbohydrates, regulating cell wall deposition and plant development. Combining multidisciplinary approaches, we study various components of the endomembrane system in the trans-Golgi Network (TGN), a site of polysaccharide trafficking and recycling of endosomal components. Made in the Golgi apparatus and assembled in the cell wall, the details of matrix polysaccharide transport have been largely unknown. This partially attributed to technical challenges in biochemically determining polysaccharide cargo in specific vesicles. To address this, we used a vesicle isolation method coupled with large scale automated glycomics and investigated the glycan contents of specific vesicle subsets. A range of diverse glycans, including pectins, xyloglucans (XyGs), and arabinogalactan proteins were identified in the isolated vesicles. This study provided a view of wall polysaccharides while they are still in the endomembrane system prior to their deposition in the apoplast. It can form the foundation of hypotheses on the biosynthesis of matrix polysaccharides in the endomembrane-system and further how they are modified after being deposited in the wall. Extensions of this study can shed light on how endomembrane glycome populations change during plant development and as a response to environmental stimuli. In plants, de novo formation of a cell plate occurs via homotypic fusion of vesicles, which develops into the new cell wall, partitioning the cytoplasm of the dividing cell. Exploiting the rapid advances in fluorescence microscopy allowing the quantitative spatio-temporal analysis of plant cytokinesis, in combination with chemical biology, we are assembling a biophysical model of cell plate formation, which will be presented. Reference: Wilkop T, et al., 2019. Plant Cell. 31(3):627-644. doi: 10.1105/tpc.18.00854.