Tubulin (porcine brain, >99% pure)

On Arrival: -70°C

Question 1: Under what conditions should I use tubulin prepared as a frozen liquid vs lyophilized powder?

Answer 1: Under most experimental conditions, we recommend using the lyophilized tubulin ((Cat. #T240) as it offers improved storage and stability compared to the frozen liquid (Cat. #T238P) which must be stored at -70°C. Some researchers favor the frozen tubulin stock for high resolution microscopy studies and crystallography work.

Question 2: Are there special instructions for thawing and aliquoting Cat. #T238P ?

Answer 2: It is recommended that the frozen liquid tubulin (Cat. #T238P) be stored at -70°C, where it is stable for 6 months from the date of purchase. To use, the protein should be rapidly thawed in a room temperature water bath, immediately transferred to ice and aliquoted into &ldquo, experiment sized&rdquo, amounts. Snap freeze aliquots in liquid nitrogen and store at -70°C. Aliquots of T238P must be snap frozen in liquid nitrogen prior to storage at -70°C. Failure to do this results in significant loss of activity. Tubulin will be active for only 1 week if stored at -40°C.

IC50 & EC50 determinations for anti-tubulin ligands.
Microtubule binding studies
Tubulin monomer binding studies
HDAC6 studies
Microtubule activated kinesin ATPase assays

The biological activity of T238P is assessed by a tubulin polymerization assay. The ability of tubulin to polymerize into microtubules is followed by observing an increase in optical density at 340nm (OD340 nm) of a tubulin solution. Under the experimental conditions defined below a 5 mg/ml tubulin solution in General tubulin buffer (Cat. # BST01) buffer plus 5% glycerol (Cat. # BST05) and 1 mM GTP (Cat. # BST06) should achieve an OD340 nm absorption reading between 0.75 - 1.10 in 30 min at 37C. The assay volume is 180 ul and assumes a spectrophotometer pathlength of 0.8 cm. See Fig 2 on the TL238 product page for an example of tubulin polymerization results.

It should be noted that tubulin minus glycerol WILL NOT polymerize in G-PEM buffer until very high tubulin concentrations (>10 mg/ml). Even at these concentrations polymerization is comparatively slow. Efficient polymerization at lower concentration of tubulin minus glycerol can be achieved by addition of a polymerization stimulating compound, e.g. paclitaxel or DMSO. See Figure 2 on the T237 product page for polymerization in the presence of glycerol and paclitaxel.