Anti-Phospho-HSF1 (Ser326) Antibody (5K398) is a Rabbit antibody targeting Phospho-HSF1 (Ser326). Anti-Phospho-HSF1 (Ser326) Antibody (5K398) can be used in WB,IHC-P,IHC-Fr,IF,FCM.

IgG

5K398

Human,Mouse (predicted:Rat)

1. Paraformaldehyde-fixed, paraffin embedded (mouse spleen); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326)) Polyclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
2. Paraformaldehyde-fixed, paraffin embedded (mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326)) Polyclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
3. Paraformaldehyde-fixed, paraffin embedded (mouse heart); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326)) Polyclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
4. Paraformaldehyde-fixed, paraffin embedded (mouse lung); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326)) Polyclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
5. Paraformaldehyde-fixed, paraffin embedded (mouse pancreas); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326)) Polyclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
6. Paraformaldehyde-fixed, paraffin embedded (human endometrial carcinoma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326)) Polyclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
7. Blank control: MCF7.
Primary Antibody (green line): Rabbit Anti-phospho-HSF1 (Ser326) antibody (TMAB-10820)
Dilution: 2 μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-AF488
Dilution: 1 μg /test.
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 90% ice-cold methanol for 20 min at-20℃. The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.
8. Paraformaldehyde-fixed, paraffin embedded (human colon carcinoma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (phospho-HSF1 (Ser326) ) Monoclonal Antibody, Unconjugated (TMAB-10820) at 1: 200 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructions and DAB staining.
9. Sample:
Lane 1: Normal human HeLa cell lysates
Lane 2: Hela cells heated at 42 ℃ for 30 minutes
Primary: Anti-phospho-HSF1 (Ser326) (TMAB-10820) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 57 kDa
Observed band size: 80 kDa

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; IF: 1:100-500; FCM: 2μg/Test

Monoclonal

KLH conjugated Synthesised phosphopeptide: human HSF1 around the phosphorylation site of Ser326

Human

HSF1

Heat shock factor protein 1

DNA-binding protein that specifically binds heat shockpromoter elements (HSE) and activates transcription. In highereukaryotes, HSF is unable to bind to the HSE unless the cells areheat shocked.