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Sample: Lane 1: Mouse Thymus tissue lysates Lane 2: Rat Thymus tissue lysates Lane 3: Human Jurkat cell lysates Primary: Anti-phospho-Rb (Ser780) (bs-1347R) at 1/1000 dilution Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 106 kDa Observed band size: 125 kDa

Paraformaldehyde-fixed, paraffin embedded (mouse colon); 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 (phospho-Rb (Ser780)) Polyclonal Antibody, Unconjugated (bs-1347R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

Independently Validated Antibody, image provided by Science Direct, badge number 029648:Formalin-fixed and paraffin embedded human breast tissue labeled with Anti-phospho-Rb/p105-Rb(Ser780) Polyclonal Antibody, Unconjugated (bs-1347R) at 1:200 followed by conjugation to the secondary antibody and DAB staining. Negative control, Human lymph node tissue, also stained postive

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-phospho-Rb/p105-Rb(Ser780) Polyclonal Antibody, Unconjugated(bs-1347R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining

MCF-7 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 (phospho-Rb (Ser780)) polyclonal Antibody, Unconjugated (bs-1347R) 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.

MCF-7 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 (phospho-Rb (Ser780)) polyclonal Antibody, Unconjugated (bs-1347R) 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.

Molt-4 cells were fixed with 4% PFA for 10min at room temperature ,permeabilized with 90% ice-cold methanol for 20 min at -20℃, and incubated in 5% BSA blocking buffer for 30 min at room temperature. Cells were then stained with phospho-Rb (Ser780) Antibody(bs-1347R)at 1:500 dilution in blocking buffer and incubated for 30 min at room temperature, washed twice with 2%BSA in PBS, followed by secondary antibody incubation for 40 min at room temperature. Acquisitions of 20,000 events were performed. Cells stained with primary antibody (green), and isotype control (orange).