4.2.7. Tubulin Polymerization Assay

The tubulin polymerization assay for tau4RDΔC, Ub2(48)tau4RD(353) and Ub2(63)tau4RD(353) was initiated by mixing in a 96-well plate 25 μM protein with tubulin (36 μM, in 100 μL total volume) in MT assembly buffer (80 mM PIPES pH 6.9, 2 mM MgCl2 and 0.5 mM EGTA) supplemented with 1 mM GTP. The plate was incubated at 37 ◦C for 4 min. Then, the polymerization was monitored by measuring the absorbance at 350 nm every 30 sec, using a Tecan Infinite M200PRO Plex plate reader (Männendorf, CH) at 37 ◦C. Each experiment was performed in triplicate.
