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Protocol for the Bottom-Up Proteomic Analysis of Mouse Spleen.

STAR protocols (2020-12-31)
Paul Dowling, Stephen Gargan, Margit Zweyer, Michael Henry, Paula Meleady, Dieter Swandulla, Kay Ohlendieck
RÉSUMÉ

This protocol describes the comparative proteomic profiling of the spleen of wild type versus mdx-4cv mouse, a model of dystrophinopathy. We detail sample preparation for bottom-up proteomic mass spectrometry experiments, including homogenization of tissue, protein concentration measurements, protein digestion, and removal of interfering chemicals. We then describe the steps for mass spectrometric analysis and bioinformatic evaluation. For complete details on the use and execution of this protocol, please refer to Dowling et al. (2020).

MATÉRIAUX
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Description du produit

Sigma-Aldrich
Acétonitrile, suitable for HPLC, gradient grade, ≥99.9%
Sigma-Aldrich
Acide trifluoroacétique, ReagentPlus®, 99%
Sigma-Aldrich
Base Trizma®, Primary Standard and Buffer, ≥99.9% (titration), crystalline
Sigma-Aldrich
Dodécylsulfate de sodium, BioReagent, suitable for electrophoresis, for molecular biology, ≥98.5% (GC)
Sigma-Aldrich
Ammonium bicarbonate, ReagentPlus®, ≥99.0%