Mouse PD-L1/B7-H1 Antibody

Detection of Mouse PD-L1/B7-H1 by Western Blot. Western blot shows lysates of RAW 264.7 mouse monocyte/macrophage cell line untreated (-) or treated (+) with 10 µg/mL LPS for 4 hours. PVDF membrane was probed with 0.5 µg/mL of Goat Anti-Mouse PD-L1/B7-H1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1019) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for PD-L1/B7-H1 at approximately 50-55 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of PD-L1/B7-H1 in RAW 264.7 Mouse Cell Line by Flow Cytometry. RAW 264.7 mouse monocyte/macrophage cell line either treated with LPS overnight (filled histogram) or untreated (open histogram) was stained with Goat Anti-Mouse PD-L1/B7-H1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1019), followed by Allophycocyanin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0108). View our protocol for Staining Membrane-associated Proteins.

Detection of PD-L1/B7-H1 in HEK293 Human Cell Line Transfected with Mouse PD-L1/B7-H1 and eGFP by Flow Cytometry. HEK293 human embryonic kidney cell line transfected with either (A) mouse PD-L1/B7-H1 or (B) irrelevant transfectants and eGFP was stained with Goat Anti-Mouse PD-L1/B7-H1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1019) followed by Allophycocyanin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0108). Quadrant markers were set based on control antibody staining (Catalog # AB-108-C). View our protocol for Staining Membrane-associated Proteins.

Detection of Mouse PD-L1 by Western Blot Pyruvate kinase isoform M2 (PKM2) is required for LPS-induced expression of PD-L1. RAW cells (0.5 × 106 cells/ml) (A), bone marrow-derived macrophages (BMDMs) (B), or bone marrow-derived dendritic cells (C) were treated with TEPP-46 at 50 µM for 1 h prior to stimulation with LPS (100 ng/ml, 24 h), lysed and assayed for expression of PD-L1 by flow cytometry [(A–C) top panels] and pdl1 mRNA by RT-PCR [(A,B) bottom panels]. Statistical analysis was performed on cells from three separate experiments. Error bars represent mean ± SEM. BMDM cells were transfected with scrambled control (SC) or anti-PKM2 siRNA (D). 24 h post-transfection, cells were stimulated with LPS (100 ng/ml) for 24 h after which expression of PKM2 (top panel) and PDL1 (middle panel) was measured by western blot. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/29081778), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of PD-L1/B7-H1 in Mouse Intestine. Formalin-fixed paraffin-embedded tissue sections of mouse intestines were probed for PD-L1 mRNA (ACD RNAScope Probe, catalog #420508; Fast Red chromogen, ACD catalog # 322750). Adjacent tissue section was processed for immunohistochemistry using goat anti-mouse PD-L1 polyclonal antibody (R&D Systems catalog # AF1019) at 3ug/mL with one-hour incubation at room temperature followed by incubation with anti-goat IgG VisUCyte HRP Polymer Antibody (Catalog # VC004) and DAB chromogen (yellow-brown). Tissue was counterstained with hematoxylin (blue). Specific staining was localized to lamina propria of villi.

Detection of Mouse Mouse PD-L1/B7-H1 Antibody by Immunohistochemistry Protumor roles for TAMs and granulocytic myeloid-derived suppressor cells are obviated in neoantigen+ PDA.(A) Immunofluorescence staining of CB+ (nAg+) or CB– (nAg–) orthotopic tumors on day 21. Arrows show CD8+ T cell contact with a macrophage. Scale bar: 50 μm. Original magnification of insets, 2.25×. The number of CD8+ T cell and TAM (F4/80+) contacts per field of view. The box plots depict the minimum and maximum values (whiskers), the upper and lower quartiles, and the median. The length of the box represents the interquartile range. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/35393950), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Mouse PD-L1/B7-H1 Antibody by Immunohistochemistry Representative images (×20 magnification) of H&E and PD-L1 IHC-stained sections (A-C): (A) Nude mouse lymph node showing normal histology (H&E) and diffuse staining for PD-L1 (mouse) with vessels and cells within the paracortical regions displaying increased staining intensity; (B) Nude mouse spleen showing normal histology (H&E) and intense membranous staining for PD-L1 (mouse) with the most intense staining at the periphery of the white pulp; (C) MDA-MB231 tumor showing a viable region (H&E necrotic regions were observed in all sections) with membranous and cytoplasmic staining for PD-L1 (human) on the tumor cells; (D) IHC quantitative analysis (staining intensity score, H-score) of PD-L1 expression levels in lymph nodes, spleen, and MDA-MB231 tumors from mouse xenografts; each bar represents the mean H-score ± standard deviation (SD; n = 3, spleen and lymph nodes; n= 6, tumors). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/31044647), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Mouse PD-L1/B7-H1 Antibody by Immunohistochemistry Representative images (×20 magnification) of H&E and PD-L1 IHC-stained sections (A-C): (A) Nude mouse lymph node showing normal histology (H&E) and diffuse staining for PD-L1 (mouse) with vessels and cells within the paracortical regions displaying increased staining intensity; (B) Nude mouse spleen showing normal histology (H&E) and intense membranous staining for PD-L1 (mouse) with the most intense staining at the periphery of the white pulp; (C) MDA-MB231 tumor showing a viable region (H&E necrotic regions were observed in all sections) with membranous and cytoplasmic staining for PD-L1 (human) on the tumor cells; (D) IHC quantitative analysis (staining intensity score, H-score) of PD-L1 expression levels in lymph nodes, spleen, and MDA-MB231 tumors from mouse xenografts; each bar represents the mean H-score ± standard deviation (SD; n = 3, spleen and lymph nodes; n= 6, tumors). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/31044647), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse Mouse PD-L1/B7-H1 Antibody by Immunohistochemistry Representative images (×20 magnification) of H&E and PD-L1 IHC-stained sections (A-C): (A) Nude mouse lymph node showing normal histology (H&E) and diffuse staining for PD-L1 (mouse) with vessels and cells within the paracortical regions displaying increased staining intensity; (B) Nude mouse spleen showing normal histology (H&E) and intense membranous staining for PD-L1 (mouse) with the most intense staining at the periphery of the white pulp; (C) MDA-MB231 tumor showing a viable region (H&E necrotic regions were observed in all sections) with membranous and cytoplasmic staining for PD-L1 (human) on the tumor cells; (D) IHC quantitative analysis (staining intensity score, H-score) of PD-L1 expression levels in lymph nodes, spleen, and MDA-MB231 tumors from mouse xenografts; each bar represents the mean H-score ± standard deviation (SD; n = 3, spleen and lymph nodes; n= 6, tumors). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/31044647), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human PD-L1 by Immunocytochemistry/ Immunofluorescence Human pancreas tissue from type 1 diabetic subjects express PD-L1. Human pancreas sections from type 1 diabetic (T1D), autoantibody positive (AA+), type 2 diabetic (T2D), and non-diabetic controls (NDB) were obtained from the Network for Pancreatic Organ Donation (nPOD) and stained for T cell markers (CD4, CD8), insulin, and PD-L1. Shown are representative islets from each group with 7–15 unique islets analyzed from three independent experiments with one patient per group. Scale bar corresponds to 20 µm. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/29844327), licensed under a CC-BY license. Not internally tested by R&D Systems.