Updated project metadata. In order to understand the molecular underpinnings of the functional and morphological heterogeneity of synapses and their regulation during development it is essential to identify the set of proteins that form these structures. Here, we prepared cortical synaptosomal fractions from juvenile and adult mice, processed them using a multidimentional protein identification technology approach, applied iTRAQ labelling and used mass spectroscopy to identify a comprehensive set of synaptic proteins and to quantitatively analyze changes in protein expression during development. We obtained a dataset tracking expression levels of 3424 proteins during development. The inclusion of biological replicates in a single mass spectrometry analysis demonstrated a high accuracy of our method. A gene ontology analysis confirmed the synaptic nature of a large fraction of identified proteins. Furthermore, developmental regulation of eight proteins identified in our analysis was confirmed independently using Western blotting. In summary, we provide a comprehensive dataset describing the developmental regulation of synaptosomal proteins, which serves as an essential foundation for the investigation of synaptic function and maturation.