Sample: Lane 1: PBMC (Mouse) Lysate at 40 ug Lane 2: Thymus (Mouse) Lysate at 40 ug Lane 3: Spleen (Mouse) Lysate at 40 ug Lane 4: Thymus (Rat) Lysate at 40 ug Primary: Anti-FoxP3 (bs-10211R) at 1/1000 dilution Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 43/45 kD Observed band size: 43/45 kD

Tissue/cell: Rat brain tissue; 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-FoxP3 Polyclonal Antibody, Unconjugated(bs-10211R) 1:500, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Tissue/cell: rat spleen; 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-FoxP3 Polyclonal Antibody, Unconjugated(bs-10211R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Blank control: Mouse spleen. Primary Antibody (green line): Rabbit Anti-FoxP3/PE Conjugated antibody (bs-10211R-PE) Dilution: 0.2μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG-PE . 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. The cells were stained with Primary Antibody for 30 min at room temperature. Acquisition of 20,000 events was performed.

Blank control: MCF7. Primary Antibody (green line): Rabbit Anti-FoxP3 antibody (bs-10211R) Dilution: 2μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG . Secondary Antibody : Goat anti-rabbit IgG-AF647 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: MCF7. Primary Antibody (green line): Rabbit Anti-FoxP3 antibody (bs-10211R) Dilution: 2μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG . Secondary Antibody : Goat anti-rabbit IgG-AF647 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 (Black line):Mouse spleen (Black). Primary Antibody (green line): Rabbit Anti-FoxP3 antibody (bs-10211R) Dilution: 3μg /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 4% PFA (10min at room temperature)and then permeabilized with 90% ice-cold methanol for 20 min at room temperature. The cells were then incubated in 5%BSA goat serum to block non-specific protein-protein interactions for 15 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.