Related Experiment Video
Updated: Aug 15, 2026
Isolation of Primary Murine Brain Microvascular Endothelial Cells
Published on: November 14, 2014
Isolation and characterization of bovine brain myelin distribution of 5'-nucleotidase
1Departament de Bioquímica i Fisiologia, Facultat de Química, Universitat de Barcelona, Spain.
Abstract:
Myelin was isolated from bovine brain by several published procedures and modifications of these procedures. High activity of the myelin marker (2',3'-cyclic nucleotide 3'-phosphohydrolase) and low activity of contaminants markers in white matter homogenates in respect to cerebral cortex showed the white matter to be better than the cerebral cortex or the whole brain for myelin isolation. A procedure is described for the preparation of purified myelin from bovine white matter which yielded a content of protein (40%), myelin marker (51%), and 5'-nucleotidase (25%) in purified myelin higher than by any used method. Acetylcholinesterase or succinate dehydrogenase was lower than 7% of its activity in the white matter homogenate, and monoamine oxidase and NADPH:cytochrome c reductase were not recovered in myelin fraction. Morphologically, myelin fraction was shown to mainly consist of multilamellar membranes of different sizes. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of myelin fraction showed a characteristic protein pattern of myelin. When our procedure was applied to frozen white matter, lower protein (32%) and myelin marker (34%) and similar 5'-nucleotidase activity (24%) were recovered in myelin, increasing its recovery in denser fractions of white matter.
Insights
This study optimized myelin isolation from bovine white matter, achieving higher purity and yield than previous methods. The refined procedure enhances the isolation of myelin for further research and analysis.
Area of Science:
- Neuroscience
- Biochemistry
- Cell Biology
Background:
- Myelin isolation is crucial for studying its structure and function.
- Existing methods have limitations in purity and yield.
- Bovine white matter is a suitable source for myelin isolation.
Purpose of the Study:
- To develop an optimized procedure for isolating highly purified myelin from bovine white matter.
- To compare the efficiency of white matter versus cerebral cortex or whole brain for myelin isolation.
- To characterize the purity and yield of myelin obtained using the novel method.
Main Methods:
- Isolation of myelin from bovine white matter using modified published procedures.
- Assay of myelin marker (2',3'-cyclic nucleotide 3'-phosphohydrolase) and contaminant markers.
- Biochemical analysis of protein content and enzyme activities.
- Morphological examination using electron microscopy.
- Protein pattern analysis using SDS-PAGE.
Main Results:
- White matter showed higher myelin marker activity and lower contaminant markers compared to cerebral cortex and whole brain.
- The optimized procedure yielded myelin with 40% protein and 51% myelin marker activity, surpassing previous methods.
- Contaminant enzyme activities (acetylcholinesterase, succinate dehydrogenase, monoamine oxidase, NADPH:cytochrome c reductase) were significantly reduced.
- Morphological and electrophoretic analyses confirmed the purity and characteristic composition of the isolated myelin.
- Application to frozen white matter yielded slightly lower but comparable results, with increased recovery in denser fractions.
Conclusions:
- Bovine white matter is a superior source for myelin isolation compared to other brain regions.
- The described procedure significantly enhances myelin purity and yield.
- The optimized method provides a reliable approach for obtaining high-quality myelin for biochemical and morphological studies.
