Phospho tacc3 antibody

Cells were lysed with lysis buffer (4 °C) containing 20 mM Tris-HCl, pH 7.4, 0.1% Triton X-100, 50 mM NaCl, 1 mM EGTA, phosphatase inhibitors 2 and 3, and a protease inhibitor mixture (Sigma). γ-tubulin was immunoprecipitated using mouse monoclonal antibody followed by addition of protein G-agarose beads. The beads were washed with lysis buffer and then boiled in SDS-PAGE sample buffer for immunoblot analysis. Membranes were developed for immunoblot using the Immobilon reagent (Millipore), followed by imaging using ChemiDoc XRS System (Bio-Rad). Ser 558-phosphorylated TACC3 was probed by rabbit polyclonal phospho-TACC3 antibody (Cell Signaling, USA).

Cells were lysed with lysis buffer (4°C) containing 50 mM HEPES, pH 7.2, 0.1% Triton X-100, 100 mM NaCl, 1 mM EGTA, 1 mM MgCl₂, phosphatase inhibitors 2 and 3, and a protease inhibitor mixture (Sigma) [46 (link)]. GFP protein was immunoprecipitated using mouse monoclonal antibody followed by the addition of protein A/G-agarose beads. GFP-coated beads (GFP-Trap) was used in the ch-TOG pull-down. The beads were washed with lysis buffer and then boiled in SDS-PAGE sample buffer for immunoblot analysis. Membranes were developed for immunoblot using the Immobilon reagent (Millipore), followed by imaging using ChemiDoc XRS System (Bio-Rad). Ser 558-phosphorylated TACC3 was probed by rabbit polyclonal phospho-TACC3 antibody (Cell Signaling, U.S.A.). For γ-tubulin and GCP4 IP, the same procedure was followed as above, using the respective antibodies.