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Sample: Lane 1: Mouse Bone tissue lysates Lane 2: Mouse Raw264.7 cell lysates Lane 3: Human Jurkat cell lysates Lane 4: Human MCF-7 cell lysates Lane 5: Human THP-1 cell lysates Primary: Anti-CCR7 (bs-1305R) at 1/1000 dilution Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 42 kDa Observed band size: 42 kDa

Paraformaldehyde-fixed, paraffin embedded (RAT lymphoid); 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 (CCR7) Polyclonal Antibody, Unconjugated (bs-1305R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Tissue/cell: human laryngocarcinoma; 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-CCR7 Polyclonal Antibody, Unconjugated(bs-1305R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (CCR7) polyclonal Antibody, Unconjugated (bs-1305R) 1:100, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.

Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (CCR7) polyclonal Antibody, Unconjugated (bs-1305R) 1:100, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.

Tissue/cell: human gastric tissue;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-CCR7 Polyclonal Antibody, Unconjugated(bs-1305R) 1:200, overnight at 4°C; The secondary antibody was Goat Anti-Rabbit IgG, FITC conjugated(bs-0295G-FITC)used at 1:200 dilution for 40 minutes at 37°C. DAPI(5ug/ml,blue,C-0033) was used to stain the cell nuclei

Blank control:THP-1. Primary Antibody (green line): Rabbit Anti-CCR7 antibody (bs-1305R) Dilution: 1μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG . Secondary Antibody : Goat anti-rabbit IgG-FITC Dilution: 0.5μg /test. Protocol The cells were fixed with 4% PFA (10min at room temperature)and then permeabilized with 0.1% PBST for 20 min at room temperature.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: Raji(blue). Primary Antibody:Rabbit Anti-CCR7 antibody(bs-1305R), Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA; Isotype Control Antibody: Rabbit IgG(orange) ,used under the same conditions ); Secondary Antibody: Goat anti-rabbit IgG-PE(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA. Protocol The cells were fixed with 2% paraformaldehyde (10 min) . Primary antibody (bs-1305R, 1μg /1x10^6 cells) were incubated for 30 min on the ice, followed by 1 X PBS containing 0.5% BSA + 10% goat serum (15 min) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/PE antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 30 min on ice. Acquisition of 20,000 events was performed.

Blank control (Black line): Mouse spleen(Black). Primary Antibody (green line): Rabbit Anti-CD4 antibody (bs-1305R-PE) Dilution: 3μg /10^6 cells; Isotype Control Antibody (orange line): Rabbit IgG-PE. 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 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.