Islets of Langerhans are critical for the homeostasis of blood glucose levels. Islets secrete antagonistic protein hormones to regulate blood sugar. Dysregulation or damage of islets can cause diabetes, a metabolic disease characterized by persistent high blood glucose levels. As islets are the primarily responsible for blood glucose interrogating the fundamental alterations helps elucidate causation and helps provide a holistic understanding of the disease. In this work, I present a systematic study of the secretome, or secreted protein content, of murine islets of Langerhans. In particular, I focused on optimizing a bottom-up proteomic approach for analysis with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Additionally, the effects of elevated glucose levels on the secretome were studied to better understand the effects of diabetes.