HeLa cell tubulin (H001)

Material
HeLa Cell tubulin is isolated from the human cervical cancer derived HeLa S3 cell line, a model system to study many aspects of tumor cell growth. Hela Cell tubulin may be used in all situations where previously bovine brain tubulin has been employed, for example drug screening, motility assays and biochemical studies including microtubule dynamics. The advantage of using this novel tubulin is that it is derived from an actively dividing cell line which often, in contrast to brain derived tubulins, more accurately portrays the situation that many researchers are trying to reconstruct in vitro.

The specificity of ligands for for a particular tubulin variant can be determined by performing comparative studies with both cancer cell and neuronal tubulins. Cytoskeleton, Inc. has advanced this concept by developing the Tubulin Ligand Index (TLI) system (patent pending). In this system, IC50 values for inhibitory compounds or EC50 values for stabilizing compounds are determined in in polymerization assays using cancer cell and neuronal tubulins. The IC50 or EC50 values for each tubulin variant are analyzed as a ratio (neuronal/cancer cell) and allow for determinations of the relative specificity for each tested compound. TLI values greater than 1.0 indicate that the particular compound is more active on cancer cell tubulin. Conversely, TLI values less than 1.0 suggest that a compound is more specific for neuronal tubulin. Table 1 summarizes data from a study comparing the specificity of several tubulin ligands using the TLI system.

Table 1. Tubulin ligand index values from studies with cancer cell and neuronal tubulins
	EC50/IC50 Value (µM)			TLI Value
Ligand	Neuronal	MCF-7	HeLa	MCF-7	HeLa
Paclitaxel Docetaxel 10-Deacetyl Taxol	0.48 0.47 3.71	0.51 0.34 4.20	1.04 0.41 30.00	0.94 1.38 0.88	0.46 1.15 0.12
Vinblastine Vincristine	1.10 1.58	1.21 n.d.	2.83 2.25	0.91 n.a.	0.39 0.70
Colchicine Nocodazole Mebendazole MF708	4.10 3.40 3.98 3.54	4.60 3.20 14.80 n.d.	3.10 3.20 25.00 1.91	0.89 1.06 0.27 n.a.	1.32 1.06 0.16 1.85

Cytoskeleton, Inc's HeLa cell tubulin protein is provided as a lyophilized powder and when reconstituted, it is in 80 mM PIPES, 1 mM MgCl_2, 1 mM EGTA and 1 mM GTP (G-PEM), pH 6.9.

HeLa cell tubulin is also available as a biotinylated conjugate for scintillation proximity assay (SPA) applications (Cat. # H003). For another human cancer cell derived tubulin, see our MCF-7 tubulin (Cat. # H005)

Purity
H001 contains >90% pure HeLa cell tubulin. Purity is determined by scanning densitometry of proteins on SDS-PAGE gels.

Biological Activity
One unit of tubulin is defined as 24 µg of purified protein (as determined by the Precision Red Advanced Protein Assay Reagent, Cat. # ADV02). HeLa cell tubulin will polymerize efficiently at a concentration of 2.0 mg/ml in the presence of 20% (v/v) glycerol. The HeLa tubulin polymerization assay has been miniaturized to 12 µl (1 unit/reaction) in order to make anti-cancer drug development feasible. Visit the page on CytoDYNAMIX screens for more information. HeLa cell tubulin is also available as a biotinylated conjugate for Scintillation Proximity Assay applications (Cat. # H003). Tubulin from a breast cancer cell line (MCF-7) is also available (Cat. # H005).

Examples of publications where this product was used:
Nunes, M., Kaplan, J., Wooters, J., Hari, M., Minnick, A. A., Jr., May, M. K., Shi, C., Musto, S., Beyer, C., Krishnamurthy, G. et al. (2005). Two photoaffinity analogues of the tripeptide, hemiasterlin, exclusively label α-tubulin. Biochemistry 44, 6844-6857.

Spittle, C., Charrasse, S., Larroque, C. and Cassimeris, L. (2000). The interaction of TOGp with microtubules and tubulin. J. Biol. Chem. 275, 20748-20753.