MCF7 cells(black) were fixed with 4% PFA for 10min at room temperature,permeabilized with 90% ice-cold methanol for 20 min at -20\u2103, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with HSF1 (S326) (37E4) Monoclonal Antibody(bsm-52166R)at 1:50 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2%BSA in PBS, followed by secondary antibody(blue) incubation for 40 min at room temperature. Acquisitions of 20,000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).

Paraformaldehyde-fixed and paraffin-embedded Mouse lung tissue incubated with HSF1 (S326) (37E4) Monoclonal Antibody (bsm-52166R) at 1:200, overnight at 4\u00b0C, followed by a conjugated secondary antibody and DAB staining. Counterstained with hematoxylin.

Paraformaldehyde-fixed and paraffin-embedded Mouse brain tissue incubated with HSF1 (S326) (37E4) Monoclonal Antibody (bsm-52166R) at 1:200, overnight at 4\u00b0C, followed by a conjugated secondary antibody and DAB staining. Counterstained with hematoxylin.

Paraformaldehyde-fixed and paraffin-embedded Mouse spleen tissue incubated with HSF1 (S326) (37E4) Monoclonal Antibody (bsm-52166R) at 1:200, overnight at 4\u00b0C, followed by a conjugated secondary antibody and DAB staining. Counterstained with hematoxylin.

Paraformaldehyde-fixed and paraffin-embedded Mouse heart tissue incubated with HSF1 (S326) (37E4) Monoclonal Antibody (bsm-52166R) at 1:200, overnight at 4\u00b0C, followed by a conjugated secondary antibody and DAB staining. Counterstained with hematoxylin.

Paraformaldehyde-fixed and paraffin-embedded Mouse pancreas tissue incubated with HSF1 (S326) (37E4) Monoclonal Antibody (bsm-52166R) at 1:200, overnight at 4\u00b0C, followed by a conjugated secondary antibody and DAB staining. Counterstained with hematoxylin.

Paraformaldehyde-fixed and paraffin-embedded Human endometrial cancer tissue incubated with HSF1 (S326) (37E4) Monoclonal Antibody (bsm-52166R) at 1:200, overnight at 4\u00b0C, followed by a conjugated secondary antibody and DAB staining. Counterstained with hematoxylin.

Paraformaldehyde-fixed, paraffin embedded Mouse lung; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with HSF1 (S326) (37E4) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, DAB staining.

Paraformaldehyde-fixed, paraffin embedded Mouse brain; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with HSF1 (S326) (37E4) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, DAB staining.

Paraformaldehyde-fixed, paraffin embedded Mouse spleen; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with HSF1 (S326) (37E4) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, DAB staining.

Paraformaldehyde-fixed, paraffin embedded Mouse heart; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with HSF1 (S326) (37E4) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, DAB staining.

Paraformaldehyde-fixed, paraffin embedded Mouse pancreas; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with HSF1 (S326) (37E4) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, DAB staining.

Paraformaldehyde-fixed, paraffin embedded Human endometrial carcinoma; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with HSF1 (S326) (37E4) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, DAB staining.

Paraformaldehyde-fixed, paraffin embedded (human colon carcinoma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (HSF1 (S326)) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Lane 1: Normal human HeLa cell lysates; Lane 2: Hela cells heated at 42 \u2103 for 30 minutes lysates probed with phospho-HSF1 (Ser326) Monoclonal Antibody, Unconjugated (bsm-52166R) at 1:1000 dilution and 4\u00b0C overnight incubation. Followed by conjugated secondary antibody incubation at 1:20000 for 60 min at 37˚C.