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In vitro tubulin polymerization assay
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Microtubule assembly was studied using the CytoDYNAMIX Screen kit (BK006P; Cytoskeleton Inc., Denver, CO, USA). Purified porcine tubulin protein (>99% purity) was resuspended in G-PEM buffer containing 80 mM PIPES, 2 mM MgCl2, 0.5 mM EGTA, 1 mM GTP (pH 6.9) and 15% glycerol in the absence or presence of indicated compounds at 4 °C. The mixture was transferred into a pre-warmed 96-well plates, and polymerization was monitored (absorbance) using a FLUOstar Omega plate reader (BMG LABTECH) at 340 nm every 1 min for 60 min at 37 °C.
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