VALIDATION IMAGES
Sample:
Lane 1: Human HeLa cell lysates
Lane 2: Human HepG2 cell lysates
Lane 3: Recombinant human HLA-C & Beta-2-MG Heterodimer protein, C-His (HEK293)
Primary: Anti-HLA-C (bsm-43093M) at 1/1000 dilution
Secondary: IRDye800CW Goat Anti-Mouse IgG at 1/20000 dilution
Predicted band size: 41 kDa
Observed band size: 45 kDa
Paraformaldehyde-fixed, paraffin embedded (human lung carcinoma); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human tonsil); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human breast carcinoma); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human liver carcinoma); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human gastric carcinoma); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human skin); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human liver); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Paraformaldehyde-fixed, paraffin embedded (human colon carcinoma); 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 (HLA-C) Monoclonal Antibody, Unconjugated (bsm-43093M) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Mouse)(sp-0024) instructionsand DAB staining.
Blank control:U266.
Primary Antibody (green line): Mouse Anti-HLA-C antibody (bsm-43093M)
Dilution: 1ug/Test;
Secondary Antibody (white blue line) : Goat anti-Mouse IgG-AF488
Dilution: 0.5ug/Test.
Isotype control(orange line):Normal Mouse IgG
Protocol
The cells were 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.