Human PD-L1/B7-H1 Antibody

Catalog #: MAB1562 Datasheet / COA / SDS

Recombinant Monoclonal Antibody

Catalog #	Availability	Size / Price	Qty
MAB1562-100
MAB1562-SP

Detection of human PD-L1/B7-H1 by Western Blot.

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Human PD-L1/B7-H1 Antibody Summary

Species Reactivity

Human

Specificity

Detects human PD-L1/B7-H1 in direct ELISAs.

Source

Recombinant Monoclonal Rabbit IgG Clone # 2340D

Purification

Protein A or G purified from cell culture supernatant

Immunogen

Mouse myeloma cell line, NS0-derived human PD-L1/B7-H1 protein
Phe19-Thr239
Accession # Q9NZQ7

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Label

Unconjugated

Applications

Recommended Concentration

Sample

Western Blot

2 µg/mL

See below

Flow Cytometry

0.25 µg/10⁶ cells

See below

Blockade of Receptor-ligand Interaction

In a functional flow cytometry test, 2.5 μg/mL of Rabbit anti-human PD-L1/B7-H1 Antibody(Catalog # MAB1562) will block the binding of Recombinant Human PD-1 His-tagged protein (Catalog # 8986-PD) to HEK293 human embryonic kidney cell line transfected with recombinant human PD-L1/B7-H1.

CyTOF-ready

Ready to be labeled using established conjugation methods. No BSA or other carrier proteins that could interfere with conjugation.

Neutralization

In a functional ELISA binding assay, 0.2-1.2 µg/mL of thisantibody will block 50% of the binding of 1 µg/mL of Recombinant HumanPD-L1/B7-H1 (Catalog # 156-B7) to immobilized Recombinant Human PD-1 Fc chimera (Catalog # 1086-PD) coated at 100 ng/mL (100 µL/well). At 2-3 µg/mL, this antibodywill block >90% of the binding.

Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.

Scientific Data

Western Blot

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Detection of human PD-L1/B7-H1 by Western Blot. Western blot shows lysates of human lung tissue. PVDF membrane was probed with 2 µg/mL of Rabbit Anti-Human PD-L1/B7-H1 Monoclonal Antibody (Catalog # MAB1562) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band was detected for PD-L1/B7-H1 at approximately 50 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Flow Cytometry

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Detection of PD-L1/B7-H1 in HEK293 Human Cell Line Transfected with Human PD-L1/B7-H1 and eGFP by Flow Cytometry. HEK293 human embryonic kidney cell line transfected with either (A) human PD-L1/B7-H1 or (B) irrelevant protein and eGFP was stained with Rabbit Anti-Human PD-L1/B7-H1 Monoclonal Antibody (Catalog # MAB1562) followed by APC-conjugated Goat anti-Rabbit IgG Secondary Antibody (F0111). Quadrant markers were set based on Rabbit IgG control antibody staining (MAB1050). View our protocol for Staining Membrane-associated Proteins.

Blockade of Receptor-ligand Interaction

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PD-1 binding to Human PD-L1/B7-H1-transfected HEK293 Human Cell Line is Blocked by Human PD-L1/B7-H1 Antibody. In a functional flow cytometry test, Recombinant Human PD-1 His-tagged protein (8986-PD, 500 ng/mL) binds to HEK293 human embryonic kidney cell line transfected with recombinant human PD-L1/B7-H1. (A) Binding is completely blocked by 2.5 µg/mL of Rabbit Anti-Human PD-L1/B7-H1 Monoclonal Antibody (MAB1562) but not by (B) Rabbit IgG Isotype Control (MAB1050). Ligand binding was detected by staining cells with Anti-His tag APC-conjugated Monoclonal Antibody (IC1050A).

Blockade of Receptor-ligand Interaction

View Larger

PD-1 binding to Human PD-L1/B7-H1-transfected HEK293 Human Cell Line is Blocked by Human PD-L1/B7-H1 Antibody. In a functional flow cytometry test, Recombinant Human PD-1 Fc Chimera Alexa Fluor® 647 protein (AFR1086, 500 ng/mL) binds to HEK293 human embryonic kidney cell line transfected with recombinant human PD-L1/B7-H1. (A) Binding is completely blocked by 25 µg/mL of Rabbit Anti-Human PD-L1/B7-H1 Monoclonal Antibody (Catalog # MAB1562) but not by (B) Rabbit IgG Isotype Control (MAB1050). View our protocol for Staining Membrane-associated Proteins.

Reconstitution Calculator

Preparation and Storage

Reconstitution

Reconstitute at 0.5 mg/mL in sterile PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.

12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.

Background: PD-L1/B7-H1

B7-H1, also known as PD-L1 and CD274, is an approximately 65 kDa transmembrane glycoprotein in the B7 family of immune regulatory molecules (1). Mature human B7-H1 consists of a 220 amino acid (aa) extracellular domain (ECD) with two immunoglobulin-like domains, a 21 aa transmembrane segment, and a 31 aa cytoplasmic domain (2). Within the ECD, human B7-H1 shares 73% and 74% aa sequence identity with mouse and rat B7-H1, respectively. Alternative splicing generates additional isoforms that either lack the first Ig-like domain or are truncated within the second Ig-like domain (3). B7-H1 is expressed on
inflammatory-activated immune cells including macrophages, T cells, and B cells (4-7), keratinocytes (8, 9), enothelial and intestinal epithelial cells (8, 10), as well as a variety of carcinomas and melanoma (11, 12). B7-H1 binds to T cell B7-1/CD80 and PD-1 (7, 8, 12-15). It suppresses T cell activation and proliferation (5, 8, 14, 16) and induces the apoptosis of activated T cells (11). It plays a role in the development of immune tolerance by promoting T cell anergy (7, 14) and enhancing regulatory T cell development (16). B7-H1 favors the development of anti-inflammatory IL-10 and IL-22 producing dendritic cells (5, 10) and inhibits the development of Th17 cells (16). In cancer, B7-H1 provides resistance to T cell mediated lysis, enhances EMT, and enhances the tumorigenic function of Th22 cells (6, 9, 12, 15).

References

Ceeraz, S. et al. (2013) Trends Immunol. 34:556.
Dong, H. et al. (1999) Nat. Med. 5:1365.
Frigola, X. et al. (2011) Clin. Cancer Res. 17:1915.
Tamura, H. et al. (2001) Blood 97:1809.
Chen, L. et al. (2007) J. Immunol. 178:6634.
Kuang, D.-M. et al. (2014) J. Clin. Invest. 124:4657.
Tsushima, F. et al. (2007) Blood 110:180.
Mazanet, M.M. and C.C.W. Hughes (2002) J. Immunol. 169:3581.
Cao, Y. et al. (2010) Cancer Res. 71:1235.
Scandiuzzi, L. et al. (2014) Cell Rep. 6:625.
Dong, H. et al. (2002) Nat. Med. 8:793.
Azuma, T. et al. (2008) Blood 111:3635.
Butte, M.J. et al. (2008) Mol. Immunol. 45:3567.
Park, J.-J. et al. (2010) Blood 116:1291.
Ritprajak, P. et al. (2010) J. Immunol. 184:4918.
Herold, M. et al. (2015) J. Immunol. 195:3584.

Long Name

Programmed Death Ligand 1

Entrez Gene IDs

29126 (Human); 60533 (Mouse); 499342 (Rat); 574058 (Porcine); 484186 (Canine); 102145573 (Cynomolgus Monkey)

Alternate Names

Avelumab; B7-H; B7H1; B7-H1; B7H1PDCD1L1; CD274 antigenMGC142294; CD274 molecule; CD274; PDCD1L1; PDCD1LG1; PDCD1LG1MGC142296; PDL1; PD-L1; PD-L1B7 homolog 1; PDL1PDCD1 ligand 1; programmed cell death 1 ligand 1; Programmed death ligand 1