Molecular characterization of myelin is a prerequisite for understanding the normal structure of the axon/myelin-unit in the healthy nervous system and abnormalities in myelin-related disorders. However, reliable molecular profiles necessitate very pure myelin membranes, in particular when considering the power of highly sensitive "omics"-data acquisition methods. Here, we recapitulate the history and recent applications of myelin purification. We then provide our laboratory protocols for the biochemical isolation of a highly pure myelin-enriched fraction from mouse brains and for its proteomic analysis. We also supply methodological modifications when investigating posttranslational modifications, RNA, or myelin from peripheral nerves. Notably, technical advancements in solubilizing myelin are beneficial for gel-based and gel-free myelin proteome analyses. We conclude this article by exemplifying the exceptional power of label-free proteomics in the mass-spectrometric quantification of myelin proteins.
Keywords: Cholesterol; Cyclic nucleotide phosphodiesterase (CNP); Demyelination; Density gradient ultracentrifugation; Lipidome/lipidomics; Mass spectrometry; Myelin; Myelin basic protein (MBP); Nerve conduction; Oligodendrocyte; Proteoform; Proteolipid protein (PLP); Proteome/proteomics; Transcriptome/transcriptomics; White matter.