To understand how cells communicate with each other, it is essential to define the cellular secretome, a collection of proteins including soluble secreted, unconventionally secreted and proteolytically-shed proteins. Quantitative methodologies to decipher the secretome are challenging, due to the requirement of large cell numbers and abundant serum proteins that interfere with the detection of low-abundant cellular secretome proteins. Here, we have use the highe perfomance secretome-protein-enrichment-with-click-sugars method (hiSPECS) for glyco-secretome analysis. We applied this method to investigate differences of hippocampal and cortical murine neurons. Additionally, we have inhibited the Alzheimer related protease BACE1 to identify potential substrates in the secretome of hippocampal neurons.