Our knowledge of the complex synaptic proteome and its relationship to physiological or pathological conditions is rapidly expanding. This has been greatly accelerated by the application of various evolving proteomic techniques, enabling more efficient protein resolution, more accurate protein identification, and more comprehensive characterization of proteins undergoing quantitative and qualitative changes. More recently, the combination of the classical subcellular fractionation techniques for the isolation of synaptosomes from the brain with the various proteomic analyses has facilitated this effort. This has resulted from the enrichment of many low abundant proteins comprising the fundamental structure and molecular machinery of brain neurotransmission and neuroplasticity. The analysis of various subproteomes obtained from the synapse, such as synaptic vesicles, synaptic membranes, presynaptic particles, synaptodendrosomes, and postsynaptic densities (PSD) holds great promise for improving our understanding of the temporal and spatial processes that coordinate synaptic proteins in closely related complexes under both normal and diseased states. This chapter will summarize a selection of recent studies that have drawn upon established and emerging proteomic technologies, along with fractionation techniques that are essential to the isolation and analysis of specific synaptic components, in an effort to understand the complexity and plasticity of the synapse proteome.