Label-Free Differential Analysis of Murine Postsynaptic Densities
This chapter provides detailed methodology for the enrichment and label-free differential analysis of postsynaptic density (PSD) proteins. Methods discussed will include tissue homogenization, subcellular fractionation, protein digestion, and label-free differential analysis after liquid chromatography–tandem mass spectrometry. When combined, these protocols facilitate the identification of receptors and signal transducers that comprise the PSD and provide an optimized workflow for the differential analysis of PSD proteomes. This strategy supports a utility for coupling fractionation with proteomics analysis to enrich for low-abundant proteins in cellular localizations that would otherwise be lost in a global tissue context.
Key wordsPostsynaptic density Label-free Differential analysis Neuroproteomics Tandem mass spectrometry Sucrose gradient centrifugation Topograph
This work was supported by the following grants: HD41697-10 (SPG), GM103533/AA016171(MJM), and AA016653/AA016171 (CCW).