This chapter discusses a brain microtubule protein preparation depleted of mitochondrial and synaptosomal components. Microtubule protein is isolated by two principal approaches: ion-exchange chromatography on DEAE-Sephadex and a recycling procedure involving warm-induced assembly and cold disassembly with intervening ultracentrifugation steps. Many microtubule purification methods apply one or both of the following methods that are capable of disrupting osmotically fragile organelles: hypotonic extraction of whole brain tissue and/or electromechanical blenders. These extraction methods contribute to destruction of brain mitochondria and synaptosomes, particularly, emulsifying effects by the lipid-rich myelinated white matter in whole brains. The sucrose method for microtubule extraction and isolation minimizes contamination. This chapter outlines a new protocol that minimizes such disruption and contamination while providing excellent yields. Additional information about the molecular characterization of sucrose purified microtubule protein is also presented in the chapter.
ASJC Scopus subject areas
- Cell Biology