Immunoflurocence analysis of human lung carcinoma cells using MLK3 (phospho-Thr277/Ser281) antibody
Blank control (Black line): A431 (Black). Primary Antibody (green line): Rabbit Anti-MLK3 antibody (orb6424), dilution: 1 µg/10 6 cells, Isotype Control Antibody (orange line): Rabbit IgG. Secondary Antibody (white blue line): Goat anti-rabbit IgG-AF647, dilution: 1 µg/Test. Protocol, The cells were fixed with 4% PFA (10 min 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 20000 events was performed.
Blank control: A431. Primary Antibody (green line): Rabbit Anti-MLK3 (Thr277 + Ser281) antibody (orb6424), dilution: 3 µg/10 6 cells, Isotype Control Antibody (orange line): Rabbit IgG. Secondary Antibody: Goat anti-rabbit IgG-AF647, dilution: 3 µg/Test. Protocol, The cells were fixed with 4% PFA (10 min 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 20000 events was performed.
Paraformaldehyde-fixed, paraffin embedded (mouse brain tissue), Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min, Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes, Blocking buffer (normal goat serum) at 37C for 30 min, Antibody incubation with (M3K11) Polyclonal Antibody, Unconjugated (orb6424) at 1:400 overnight at 4C, followed by a conjugated secondary for 20 minutes and DAB staining.
Paraformaldehyde-fixed, paraffin embedded (Mouse brain), Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min, Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes, Blocking buffer (normal goat serum) at 37C for 30 min, Antibody incubation with (Phospho-MLK3 (Thr277 + Ser281)) Polyclonal Antibody, Unconjugated (orb6424) at 1:500 overnight at 4C, followed by a conjugated secondary for 20 minutes and DAB staining.
Paraformaldehyde-fixed, paraffin embedded (rat liver), Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min, Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes, Blocking buffer (normal goat serum) at 37C for 30 min, Antibody incubation with (Phospho-MLK3 (Thr277 + Ser281)) Polyclonal Antibody, Unconjugated (orb6424) at 1:400 overnight at 4C, followed by a conjugated secondary for 20 minutes and DAB staining.
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