Ix81 inverse microscope

MCF-7 cells and MCF-7/ADR cells were cultured on coverslips. Then, the cells were fixed, permeabilized in 0.1% Triton X-100 and blocked in 1% BSA. The coverslips were incubated with anti-N-cadherin (ab76057, Abcam) or anti-Slug (ab27568, Abcam) at 4°C overnight. Alexa Fluor^® 488 Conjugate Secondary Anti-Rabbit (#4412, Cell Signaling) was used to detect N-cadherin, while Alexa Fluor^® 555 Conjugate Secondary Anti-Rabbit (#4413, Cell Signaling) was used to detect Slug. The coverslips were incubated with the secondary antibodies for 1 h at room temperature in the dark. Nuclei were stained using Prolong^® Gold Antifade Reagent with DAPI (#8961, Cell Signaling). Immunofluorescent images were obtained using an Olympus IX81 inverse microscope (Tokyo, Japan). Each experiment was performed in triplicate.

SiHa cells after transfection were cultured on coverslips. Then, the cells were fixed, permeabilized in 0.1% Triton X-100 and blocked in 1% BSA. The coverslips were incubated with primary antibodies against E-cadherin (#3195, Cell Signaling) and N-cadherin (#13116, Cell Signaling) respectively at 4°C overnight. Secondary Anti-Rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 555 Conjugate) (#4413, Cell Signaling) was used to detect E-cadherin, while Anti-Rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) (#4412, Cell Signaling) was used to detect N-cadherin. The coverslips were incubated with the secondary antibodies for 1 hour at room temperature in the dark. Nuclei were stained using Prolong® Gold Antifade Reagent with DAPI (#8961, Cell Signaling). Immunofluorescent images were obtained using an Olympus IX81 inverse microscope (Tokyo, Japan). Each experiment was performed in triplicate.