Anti-PDL1 CD274 Monoclonal Antibody [6H10]

Overexpression Validation of PD-L1 in 293 Cells
Loading: 15 μg of lysates per lane. Antibodies: M00109-3 (A, 0.25 μg/mL; B, 0.5 μg/mL), 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-mouse IgG HRP conjugate at 1:5000 dilution.

Independent Antibody Validation (IAV) via Protein Expression Profile in Cell Lines
Loading: 15 μg of lysates per lane. Antibodies: 4059 (2 μg/mL), M00109-3 (2 μg/mL), and beta-actin (1 μg/mL), 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-rabbit and or anti-mouse IgG HRP conjugate at 1:10000 and 1:5000 dilution, respectively.

Validation with PD-L1 siRNA Knockdown in HeLa Cells
HeLa cells were transfected with control siRNAs (lane 1) or PD-L1 siRNAs (lane 2) Loading: 10 μg of HeLa whole cell lysates per lane. Antibodies: M00109-3 (2 μg/mL) and GAPDH (3783, 0.02 μg/mL), 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-mouse IgG HRP conjugate at 1:5000 dilution.

Western Blot Validation of PD-L1 in Raji Cells
Loading: Lysates/proteins at 15 μg per lane. Antibodies: M00109-3 (4 μg/mL). 1 h incubation at RT in 5% NFDM/TBST. Secondary: Goat anti-mouse IgG HRP conjugate at 1:10000 dilution.

Immunofluorescence Validation of PD-L1 in Transfected 293 Cells
Immunofluorescent analysis of 4% paraformaldehyde-fixed PD-L1 transfected 293 cells labeling PD-L1 with M00109-3 at 2 μg/mL, followed by goat anti-mouse IgG secondary antibody at 1/500 dilution (red) and DAPI staining (blue).

Immunofluorescence Validation of PD-L1 in Human Stomach Carcinoma Tissue
Immunofluorescent analysis of 4% paraformaldehyde-fixed human stomach carcinoma tissue labeling PD-L1 with M00109-3 at 2 μg/mL, followed by goat anti-mouse IgG secondary antibody at 1/500 dilution (red) and DAPI staining (blue).

Immunofluorescence Validation of PD-L1 in Human Tonsil Tissue
Immunofluorescent analysis of 4% paraformaldehyde-fixed human tonsil tissue labeling PD-L1 with M00109-3 at 2 μg/mL, followed by goat anti-mouse IgG secondary antibody at 1/500 dilution (red) and DAPI staining (blue).

Immunohistochemistry Validation of PD-L1 in Human Stomach Carcinoma Tissue
Immunohistochemical analysis of paraffin-embedded human stomach carcinoma tissue using anti-PD-L1 antibody (M00109-3) at 5 μg/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4˚C. A goat anti-mouse IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin. Lower left: Use mouse IgG antibody at 1 μg/ml as control.

Immunohistochemistry Validation of PD-L1 in Human Tonsil Carcinoma Tissue
Immunohistochemical analysis of paraffin-embedded human tonsil tissue using anti-PD-L1 antibody (M00109-3) at 5 μg/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4˚C. A goat anti-mouse IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin. Lower left: Use mouse IgG antibody at 1 μg/ml as control.

Immunocytochemistry Validation of PD-L1 in PD-L1 Overexpressed 293 Cells
Immunocytochemical analysis of PD-L1 transfected 293 cells using anti- PD-L1 antibody (M00109-3) at 1 μg/mL. Cells was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4˚ C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin. Lower left: Use mouse IgG antibody at 1 μg/mL as control.