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Methods

Quantification of the synaptosomal proteome of the rat cerebellum during post-natal development

¹ Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA; ² Genomics Institute of the Novartis Research Foundation, San Diego, California 92121, USA

Large-scale proteomic analysis of the mammalian brain has been successfully performed with mass spectrometry techniques, such as Multidimensional Protein Identification Technology (MudPIT), to identify hundreds to thousands of proteins. Strategies to efficiently quantify protein expression levels in the brain in a large-scale fashion, however, are lacking. Here, we demonstrate a novel quantification strategy for brain proteomics called SILAM (Stable Isotope Labeling in Mammals). We utilized a ¹⁵N metabolically labeled rat brain as an internal standard to perform quantitative MudPIT analysis on the synaptosomal fraction of the cerebellum during post-natal development. We quantified the protein expression level of 1138 proteins in four developmental time points, and 196 protein alterations were determined to be statistically significant. Over 50% of the developmental changes observed have been previously reported using other protein quantification techniques, and we also identified proteins as potential novel regulators of neurodevelopment. We report the first large-scale proteomic analysis of synaptic development in the cerebellum, and we demonstrate a useful quantitative strategy for studying animal models of neurological disease.

E-mail jyates{at}scripps.edu; fax (858) 784-8883.

[Supplemental material is available online at www.genome.org.]

Article published online before print. Article and publication date are at http://www.genome.org/cgi/doi/10.1101/gr.6375007

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Related Protocol

Stable Isotope Labeling of Mammals (SILAM)

Daniel B. McClatchy and John R. Yates, III
CSH Protocols 2008 2008: pdb.prot4940. [Abstract] [Full Text]

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