Tissue/cell: human pancreas carcinoma; 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-Sna1l Polyclonal Antibody, Unconjugated(bs-1371R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Tissue/cell: human cervical carcinoma; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: 0.01M TBS (pH 7.5 ), 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-Sna1l Polyclonal Antibody, Unconjugated(bs-1371R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Tissue/cell: human laryngocarcinoma;4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
Incubation: Anti-Sna1l Polyclonal Antibody, Unconjugated(bs-1371R) 1:200, overnight at 4°C; The secondary antibody was Goat Anti-Rabbit IgG, Cy3 conjugated(bs-0295G-Cy3)used at 1:200 dilution for 40 minutes at 37°C.

Blank control: HepG2.
Primary Antibody (green line): Rabbit Anti-SNAIL antibody (bs-1371R)
Dilution: 1μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG .
Secondary Antibody : Goat anti-rabbit IgG-AF488
Dilution: 1μg /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.

Blank control (blue line): Hela (blue).
Primary Antibody (green line): Rabbit Anti-SNAIL antibody (bs-1371R)
Dilution: 1μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG .
Secondary Antibody (white blue line): Goat anti-rabbit IgG-PE
Dilution: 1μg /test.
Protocol
The cells were fixed with 70% methanol (Overnight at 4℃) and then permeabilized with 90% ice-cold methanol for 20 min at -20℃. Cells stained with Primary Antibody for 30 min at room temperature. The cells were then incubated in 1 X PBS/2%BSA/10% goat serum to block non-specific protein-protein interactions followed by the antibody for 15 min at room temperature. The secondary antibody used for 40 min at room temperature. Acquisition of 20,000 events was performed.