Sample: Lung (Mouse) Lysate at 30 ug
Primary: Anti- PTEN (SL0748R) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution
Predicted band size: 44 kD
Observed band size: 51 kD
Sample: Brain(Mouse) Lysate at 30 ug
Primary: Anti- PTEN (SL0748R) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution
Predicted band size: 44 kD
Observed band size: 51 kD
Tissue/cell: Human nasopharyngeal 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,SLC0005) at 37∩ for 20 min;
Incubation: Anti-PTEN Polyclonal Antibody, Unconjugated(SL0748R) 1:200, overnight at 4∑C, followed by conjugation to the secondary antibody(SP-0023) and DAB(SLC0010) staining
Hela cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, SLC0005) at 37°C for 20 min; Antibody incubation with (PTEN) polyclonal Antibody, Unconjugated (SL0748R) 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.
Blank control: Hela.
Primary Antibody (green line): Rabbit Anti-PTEN antibody (SL0748R)
Dilution: 1μg /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 (blue line): A431 cells (blue).
Primary Antibody (green line): Rabbit Anti-PTEN antibody (SL0748R)
Dilution: 1μg /10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG .
Secondary Antibody (white blue line): Goat anti-rabbit IgG-FITC
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.
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