Immunofluorescence-based Analysis of HSF1 Distribution

The distribution of HSF1 was assessed by immunofluorescence as described previously [26 (link)]. Briefly, the cells were fixed with 4% paraformaldehyde (G1101-500ML, Servicebio, Wuhan, China) for 15 min, permeabilized with 1% Triton X-100 (G5060-100ML, Servicebio, Wuhan, China) for 10 min, and incubated overnight at 4 °C with anti-HSF1 primary antibody (A13765, 1:100, Abclonal, Wuhan, China). Next, the cells were incubated with Cy3-labeled goat anti-rabbit antibody (BA1032, Boster, Wuhan, China) for 1 h and DAPI (AR1177, Boster, Wuhan, China) for 15 min at room temperature. The distribution of HSF1 was then observed by fluorescence microscopy (IX73-A12FL/PH, OLYMPUS, Japan).