293T (H) cells were treated with or without PMA (50 ng/mL) for 48 h, IFN-γ (50 U/mL) for 24 h, 25 μg total protein per lane of cell lysates (see on figure) probed with HLA F monoclonal antibody, unconjugated (bsm-62338R) at 1:1000 dilution and 4°C overnight incubation. Followed by conjugated secondary antibody incubation at r.t. for 60 min.

Paraformaldehyde-fixed, paraffin embedded Human Tonsil; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

Paraformaldehyde-fixed, paraffin embedded Human Placenta; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

Paraformaldehyde-fixed, paraffin embedded Human Lymph; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

Paraformaldehyde-fixed, paraffin embedded Human Esophageal Cancer; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

Paraformaldehyde-fixed, paraffin embedded Human Colon Cancer; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

Paraformaldehyde-fixed, paraffin embedded Human Cervical Cancer; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

Paraformaldehyde-fixed, paraffin embedded Human Lung Cancer; Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; The section was incubated with HLA F Monoclonal Antibody, Unconjugated (bsm-62338R) at 1:200 overnight at 4°C, followed by conjugation to the bs-0295G-HRP and DAB (C-0010) staining.

4% Paraformaldehyde-fixed Raji (H) cell; Antibody incubation with (HLA F) monoclonal Antibody, unconjugated (bsm-62338R) 1:100, 90 min at 37°C; followed by conjugated Goat Anti-Rabbit IgG antibody (green, bs-60295G-BF488) at 37°C for 90 min, DAPI (blue, C02-04002) was used to stain the cell nuclei. PBS instead of the primary antibody was used as the blank control.

The Raji (H) cells were incubated in 5%BSA to block non-specific protein-protein interactions (30 min at r.t.) , followed by secondary antibody incubation for 40 min at room temperature. Primary Antibody (green): Rabbit Anti-HLA F antibody (bsm-62338R,1:100); Isotype Control (orange): Rabbit IgG (bs-0295P). Blank control (black): PBS. Acquisition of 20,000 events was performed.