Tubulin Isolation
Tubulin Isolation - Related Videos
02:40
Second Harmonic Generation Imaging in a Rat Model to Study Tubulin Defects
0 Views •
2025
Source: Piazza, V., et al., Label-Free Non-Linear Optics for the Study of Tubulin-Dependent Defects in Central Myelin. J. Vis. Exp. (2023)This video demonstrates the procedure for imaging microtubule abnormalities in a rat brain tissue slice using a two-photon excitation microscope. It employs second harmonic generation (SHG) signals to detect tubulin defects and reduced myelin production.
06:30
Optimizing Tubulin Yield from Porcine Brain Tissue
0 Views •
2024
This protocol describes a technique for the high-yield isolation of tubulin from the porcine brain optimized for small-scale instrumentation. The isolation procedures are complemented by procedures for determining tubulin polymerization activity in vitro using co-sedimentation assays and transmission electron microscopy.
08:07
Label-Free Non-Linear Optics for the Study of Tubulin-Dependent Defects in Central Myelin
0 Views •
2023
In this article, we present a protocol to detect microtubule-loaded oligodendrocytes in a model of tubulinopathy through a simple, innovative second harmonic generation microscopy approach.
07:54
Purification of Tubulin with Controlled Posttranslational Modifications and Isotypes from Limited Sources by Polymerization-Depolymerization Cycles
0 Views •
Cited by 6 •
2020
This protocol describes tubulin purification from small/medium-scale sources such as cultured cells or single mouse brains, using polymerization and depolymerization cycles. The purified tubulin is enriched in specific isotypes or has specific posttranslational modifications and can be used in in vitro reconstitution assays to study microtubule dynamics and interactions.
07:21
Quantitative Microtubule Fractionation Technique to Separate Stable Microtubules, Labile Microtubules, and Free Tubulin in Mouse Tissues
0 Views •
2023
Microtubules, which are tubulin polymers, play a crucial role as a cytoskeleton component in eukaryotic cells and are known for their dynamic instability. This study developed a method for fractionating microtubules to separate them into stable microtubules, labile microtubules, and free tubulin to evaluate the stability of microtubules in various mouse tissues.
FAQs
Purification takes advantage of tubulin’s reversible response to GTP and environmental conditions. When polymerization is favored, tubulin joins microtubules while many other cellular proteins remain soluble, allowing the polymerized fraction to be separated. Reversing those conditions releases tubulin again. This selective cycling converts a dynamic assembly property into a biochemical enrichment strategy.
Cooling or changing the reaction conditions disassembles collected microtubules and recovers soluble tubulin. This step resets the material for another assembly cycle rather than serving only as a cleanup stage. Repeating assembly and disassembly progressively enriches tubulin because each cycle separates polymerization-competent tubulin from proteins that do not follow the same behavior.
Taxol serves a different purpose from GTP-driven assembly. Rather than creating the reversible enrichment cycle, it can stabilize polymerized material so microtubules remain intact during purification. This is useful when maintaining the assembled fraction matters, whereas cooling or altered conditions provide the contrasting route for disassembly and tubulin recovery.
A basic workflow begins with a cellular extract, followed by conditions that promote tubulin assembly in the presence of GTP. The resulting microtubules are separated from soluble proteins and then disassembled by cooling or changing the reaction conditions. Recovered tubulin can undergo additional assembly and disassembly cycles to increase enrichment before downstream studies.
Successful enrichment depends on controlling the conditions that switch tubulin between soluble and polymerized states. GTP and suitable assembly conditions favor microtubule formation, whereas cooling or altered conditions favor disassembly. Taxol can preserve polymerized material when needed. Managing these opposing states determines whether the procedure efficiently separates tubulin from the cellular extract.
Isolated tubulin supports examination of cytoskeletal dynamics and intracellular transport, as well as studies of cell division and protein interactions. It also enables investigation of how microtubule-targeting drugs act. By providing tubulin separated from the original cellular extract, the preparation supports focused structural and functional studies of these biochemical processes.