Subcellular fractionation of presynaptic and postsynaptic proteins in forebrain neurons.

<p>(<b>A</b>) Schematic illustration of the enrichment process, highlighting the key fractions examined in this study (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0042327#s2" target="_blank">Methods</a> for details). The PNS fraction obtained from the homogenate is subjected to a series of centrifugation steps to isolate pre- and postsynaptic plasma membranes (SPM), including layering on a sucrose gradient, and finally extracting with detergent to obtain the postsynaptic densities (PSD). (<b>B</b>) Representative immunoblots show NR1, PSD-95, and synaptophysin (Syp) expression in fractions from P16 mouse forebrain, with 5 or 10 µg total protein loaded in each lane as indicated. (<b>C</b>) Quantification of fraction fold-enrichment made with sample values normalized to PNS fractions. Error bars represent SEM. PNS, postnuclear supernatant; CYT, cytosol; LM, light membranes; P2, crude synaptosomes; P3, lysed synaptic membranes; SPM, purified synaptic plasma membranes; TSF, Triton-soluble fraction; PSD, postsynaptic densities.