Celllight reagents bacmam 2

Mouse monoclonal anti- antibody was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). Rabbit anti-tubulin α antibody was from Sigma-Aldrich (Oakville, Ontario). All secondary antibodies conjugated with FITC and TRITC were obtained from Life Technologies Inc (Burlington, ON, Canada). Mammalian Protein Extraction Reagent (M-Per) was purchased from Thermo Fisher Scientific Inc. (Rockford, IL). Vybrant MTT Cell Proliferation Assay Kit and GFP tagged α-tubulin with the CellLight^® Reagents BacMam 2.0 were from Invitrogen (Grand Island, NY). Unless otherwise specified, all other chemicals were purchased from Sigma-Aldrich.

Live imaging was used to study docetaxel induced M-phase arrest and microtubule dynamics. As described previously [27 ], MCF-7_CC and MCF-7_TXT cells were cultured on 35mm poly-L-lysine-coated coverslips (Fisher) overnight. To assay docetaxel-induced M-phase arrest, cells were incubated with DMEM containing 250 ng/ml Hoechst 33342 (Calbiochem) for 10 min to stain DNA. Then, coverslips were mounted on a sample holder and incubated with DMEM without phenol red, supplemented with 10% FBS with or without docetaxel. To assay microtubule dynamics, cells were transfected with GFP tagged α-tubulin with the CellLight^® Reagents BacMam 2.0 (Invitrogen, Grand Island, NY) for 24 h according to the Manufacturer’s instructions. Coverslips were then mounted on a sample holder and incubated with DMEM without phenol red, supplemented with 10% FBS either with or without docetaxel as indicated for 1 h.
Experiments were performed in a chamber maintained at 37°C and 5% CO₂. Fluorescence images were acquired at the indicated times using a Delta Vision microscopic system. Delta Vision SoftWoRx software was used to deconvolve images and generate movies.

Mouse monoclonal anti-ABCG2 (BXP-21), rabbit polyclonal anti-MDR (ABCB1) (H-241), and rabbit polyclonal anti-MRP1 (ABCC1) (E-19) antibodies were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). Rabbit anti-tubulin β1 and β3, and mouse monoclonal anti-tubulin β2 and β4 antibodies were purchased from Abcam (Toronto, ON). All secondary antibodies conjugated with FITC and TRITC were from Life Technologies Inc (Burlington, ON). Mammalian Protein Extraction Reagent (M-Per) was purchased from Thermo Fisher Scientific Inc. (Rockford, IL USA). Vybrant MTT Cell Proliferation Assay Kit and GFP tagged α-tubulin with the CellLight® Reagents BacMam 2.0 were from Invitrogen (Grand Island, NY). Unless otherwise specified, all chemicals were purchased from Sigma-Aldrich.