Isolation of microtubules and microtubule proteins.
This unit describes various protocols for the isolation and purification of the main constituents of microtubules, chiefly alpha- and beta-tubulin, and the most significant microtubule associated proteins (MAPs), specifically MAP1A, MAP1B, MAP2, and tau. We include a classical isolation method for soluble tubulin heterodimer as the first basic purification protocol. In addition, we show how to analyze the tubulin and MAPs obtained after a phosphocellulose chromatography purification procedure. This unit also details a powerful and simple method to determine the native state of the purified tubulin based on one-dimensional electrophoresis under nondenaturing conditions (UNIT 6.5). The last protocol describes the application of a new technique that allows visualizing the quality of polymerized microtubules based on atomic force microscopy (AFM).
Avila
J
J
Centro de Biología Molecular, Facultad de Ciencias, Universidad Autónoma de Madrid, Cantoblanco, Madrid, Spain.
Soares
H
H
Fanarraga
M L
ML
Zabala
J C
JC
D002845:Chromatography; D004586:Electrophoresis / Q000379:methods*; D018625:Microscopy, Atomic Force; D008868:Microtubule Proteins / Q000032:analysis / Q000302:isolation & purification* / Q000648:ultrastructure; D008870:Microtubules / Q000737:chemistry* / Q000648:ultrastructure; D017239:Paclitaxel / Q000032:analysis
D008868:Microtubule Proteins; D017239:Paclitaxel