Texas red conjugated goat anti rabbit igg

For immunocytochemistry, cells were cultured in appropriate 8-well chamber slides. After culture, CCF-STTG1 or NMW7 cells were fixed with 4% paraformaldehyde in phosphate-buffered saline (PBS) for 15 min and blocked with 5% normal goat serum and 0.1% Triton X-100 in PBS for 30 min. Then, anti-His6 IgG (mouse, 1:50; Roche, Basel, Switzerland) or anti-CST3 IgG (rabbit, 1:200; Abcam) was added to the wells and incubated overnight at 4 °C. Immunoreactive proteins were detected with Texas Red-conjugated goat anti-rabbit IgG (1:200; Santa Cruz Biotechnology, Santa Cruz, CA, USA) or FITC-conjugated goat anti-mouse IgG (1:200; Santa Cruz Biotechnology). The fluorescence signals were visualized using a fluorescence microscope equipped with filters for individual fluorophores (FITC emission filter wavelength 525 nm, Texas red emission filter wavelength 620 nm, Hoechst emission filter wavelength 460 nm) (ECLIPSE E600, NIKON, Tokyo, Japan).

7,12-Dimethylbenz[α]anthracene (DMBA), mitomycin C, mithramycin A, 4-hydroxyestradiol, 2-hydroxyestradiol, and charcoal-stripped FBS were purchased from Sigma (St. Louis, MO, USA). 2,2′,4,6′-Tetramethoxystilbene (TMS) was kindly provided by Dr. Sanghee Kim (Seoul National University, Seoul, Korea). Rabbit polyclonal antibody for E-cadherin was purchased from Millipore (Bedford, MA, USA). M-MLV reverse transcriptase and RNase inhibitor were purchased from Promega (Madison, WI, USA). Ex Taq Polymerase was obtained from TaKaRa Bio (Shiga, Japan). SYBR green was purchased from QIAGEN (Hilden, Germany). Rabbit polyclonal antibodies for CYP1B1, Sp1, β-catenin, E-cadherin, cyclin D1, vimentin, SNAI1, and GAPDH; mouse monoclonal antibody for ZEB2 and c-Myc; Texas Red-conjugated goat anti-rabbit IgG; and UltraCruz^TM Mounting Medium were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). HRP-conjugated goat anti-rabbit IgG and DyLight^® 594-conjugated goat anti-mouse were obtained from Bethyl (Montgomery, TX, USA) and mouse monoclonal antibody for PCNA was purchased from Cell Signaling Technology (Beverly, MA, USA). Other chemicals and reagents were of the highest quality commercially available.