Vimentin Mouse Monoclonal Antibody

Blank control: Hela. Primary Antibody (green line): Mouse Anti-Vimentin antibody (orb499662), dilution: 1 ug/Test, Secondary Antibody: Goat anti-Mouse IgG-FITC, dilution: 0.5 ug/Test. Protocol, The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 90% ice-cold methanol for 20 min at -20°C. 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.

Hela cell, 4% Paraformaldehyde-fixed, Triton X-100 at room temperature for 20 min, Blocking buffer (normal goat serum) at 37°C for 20 min, Antibody incubation with (Vimentin) monoclonal Antibody, Unconjugated (orb499662) 1:100, 90 minutes at 37°C, followed by a conjugated Goat Anti-Mouse IgG antibody at 37°C for 90 minutes, DAPI (blue) was used to stain the cell nuclei.

Paraformaldehyde-fixed, paraffin embedded (Human cervical cancer), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (ascites of orb499662 3B7) at 1:2000 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructions and DAB staining.

Paraformaldehyde-fixed, paraffin embedded (human cervical carcinoma), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (human colon), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (human endometrial carcinoma), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (Human kidney), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (Human lung cancer), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (ascites of orb499662 3B7) at 1:2000 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructions and DAB staining.

Paraformaldehyde-fixed, paraffin embedded (human lung carcinoma), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (human tonsil), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Paraformaldehyde-fixed, paraffin embedded (Human uterus), 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 37°C for 30 min, Antibody incubation with (Vimentin) Monoclonal Antibody, Unconjugated (orb499662) at 1:200 overnight at 4°C, followed by operating according to SP Kit (Mouse) instructionsand DAB staining.

Sample: Hela (Human) Cell Lysate at 30 ug, Hela KO Vimentin (Human) Cell Lysate at 30 ug, Primary: Anti-Vimentin (orb499662) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution, Predicted band size: 53 kD, Observed band size: 58 kD.

Sample: Raji Cell Lysate at 40 ug, U87MG Cell Lysate at 40 ug, Primary: Anti-Vimentin (orb499662) at 1/1000 dilution, Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution, Predicted band size: 53kD, Observed band size: 58kD.