Sample:Hela Cell lysate at 30ug;
Primary: Anti-S100A4 (bs-3759R) at 1:300 dilution;
Secondary: HRP conjugated Goat-Anti-Rabbit IgG(bs-0295G-HRP) at 1: 5000 dilution;
Predicted band size :12 kD
Observed band size :15 kD

Sample: Raji Cell lysate at 30ug;
Primary: Anti-S100A4 (bs-3759R) at 1:300 dilution;
Secondary: HRP conjugated Goat-Anti-Rabbit IgG(bse-0295G-HRP) at 1: 5000 dilution;
Predicted band size :12 kD
Observed band size : 12 kD

Paraformaldehyde-fixed, paraffin embedded (mouse bladder); 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°C for 30min; Antibody incubation with (S100A4) Polyclonal Antibody, Unconjugated (bs-3759R ) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (Rat bladder); 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°C for 30min; Antibody incubation with (S100A4) Polyclonal Antibody, Unconjugated (bs-3759R ) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Tissue/cell: human lung cancer; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
Incubation: Anti-S100A4 Polyclonal Antibody, Unconjugated(bs-3759R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Paraformaldehyde-fixed, paraffin embedded (Human breast 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°C for 30min; Antibody incubation with (S100A4) Polyclonal Antibody, Unconjugated (bs-3759R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Blank control: THP-1.
Primary Antibody (green line): Rabbit Anti-S100A4 antibody (bs-3759R)
Dilution: 1ug/Test;
Secondary Antibody : Goat anti-rabbit IgG-FITC
Dilution: 0.5ug/Test.
Protocol
The cells were fixed with 4% PFA (10min 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.