Cy3 labeled goat anti mouse igg

Endothelial activation marker endothelin-1 (ET-1) expression in the PMVECs was evaluated by immunofluorescence assay. As described previously, the PMVECs were fixed, permeabilized, and then incubated with anti-mouse ET-1 (1:100; Bioss, Beijing, China) at 4℃ overnight. Next, the cells were incubated with Cy3-labeled goat anti-mouse IgG (1:200; Abcam, Cambridge, UK) at room temperature for one hour. The nuclei were counterstained by DAPI (Aladdin, Shanghai, China). The images were captured using a fluorescence microscope (Olympus, Japan).

Cells were placed on glass slides, fixed with 4% paraformaldehyde for 15 min, permeabilized with 0.1% Triton X-100 for 30 min, and then blocked with 5% goat serum for 15 min at room temperature. Subsequently, cells were incubated with diluted primary antibody against Nrf2 (1:200; cat. no. ab89443; Abcam, Hong Kong) at 4°C overnight, followed by incubation with Cy3-labeled goat antimouse IgG (1:2,000; cat. no. 33208ES60; Yeason, Shanghai, China) for 60 min at room temperature. Nuclei were re-stained with DAPI, and the results of the staining process were observed under a fluorescence microscope.