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Isolation of Synapse Subdomains by Subcellular Fractionation Using Sucrose Density Gradient Centrifugation

  • Tatsuo SuzukiEmail author
Protocol
Part of the Neuromethods book series (NM, volume 57)

Abstract

A protocol presents a purification of postsynaptic density (PSD), from rat brain by subcellular fractionation using solubilization of membrane with Triton X-100 and sucrose density centrifugation. The protocol also includes purification of other synapse subdomains such as synaptosome, synaptic plasma membrane, P1 (nuclei and cell debris), P2 (crude mitochondria fraction), S3 (soluble fraction), and P3 (microsomal fraction). The method presented in this text is the one established by Siekevitz group. The PSDs obtained by this method are mainly excitatory type I PSDs. The method has been widely used and is useful for biochemical analyses such as identification of proteins associated with these subdomains by proteomics methods and western blotting, and morphological analyses at the electron microscopic level.

Key words

Synaptosome Synaptic plasma membrane Postsynaptic density Subcellular fractionation Detergent-insoluble cytoskeleton Detergent-insoluble membrane 

Notes

Acknowledgments

The author learned the method of PSD purification in the Philip Siekevitz laboratory, Rockefeller University, New York. The author heartily thanks Dr. Philip Siekevitz and Marie LeDoux for their instruction.

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.Department of Neuroplasticity, Institute on Aging and AdaptationShinshu University Graduate School of MedicineMatsumotoJapan

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