Pancreatic -cells in the islets of Langerhans are key to maintaining glucose homeostasis, by secreting the peptide hormone insulin. Insulin is packaged within vesicles named insulin secretory granules (ISGs), that have recently been considered to have intrinsic structures and proteins that regulate insulin granule maturation, trafficking, and secretion. Previously, studies have identified a handful of novel ISG-associated proteins using different separation techniques. Here, this study combines an optimized ISG isolation technique and mass spectrometry-based proteomics, with an unbiased and targeted machine learning approach to uncover 211 ISG-associated proteins. Five of these proteins have not been previously ISG-associated: Syntaxin-7, Synaptophysin, Synaptotagmin-13, Zinc transporter ZIP8 and Scamp3. We validate the role for one of these novel ISG-associated proteins (Scamp3) in regulating insulin storage and secretion from -cells for the first time. These data provide the basis for future investigation into beta cell biology and the regulation of insulin secretion.