Human PD-L1 protein, Fc-fusion, biotinylated, recombinant

Description

PD-L1 (programmed cell death 1 ligand 1) or CD274 is a 290-amino acid single pass type I transmembrane protein of the Ig (immunoglobulin) superfamily.  It contains one Ig V-like and one Ig C-like domains in the extracellular region, and a 30-amino acid cytoplasmic tail. It was originally cloned as a homolog of B7 (termed B7-H1). It is 20% and 15% identical to B7-1 (CD80) and B7-2 (CD86), respectively.  It was also cloned as a ligand (termed PDCD1L1 or PD-L1) that binds to PD-1 (CD279) and B7-1 (CD80), but not to other B7 family members, such as CTLA4 (CD152), CD28, or ICOS (CD278).  PD-L1 or CD274 is involved in the T cell costimulatory signal, essential for T-cell proliferation and production of IL10 and IFNγ. Interaction with PD-1, which has a cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM), inhibits T-cell proliferation and cytokine production.  PD-L1 is up-regulated on T- and B-cells, dendritic cells, keratinocytes and monocytes after LPS and IFNγ activation or by surface Ig cross-linking. It is expressed in many types of freshly isolated human cancers but not in most normal tissues.  Blockade of PD-L1 upregulates IL12 expression and enhances antitumor immunity in mice bearing ovarian tumors.  Therefore, PD-L1 has been proposed as a target for cancer immunotherapy through pre-activated T cells that could be enhanced by blockade of PD-L1-induced inhibitory signaling. 

 

 

Gene Symbol: PD-L1; CD274; B7-H1; B7H1;  PDL1;  PDCD1L1; PDCD1LG1

 

NCBI Gene ID: 29126

 

Uniprot Entry: Q9NZQ7

 

Construct Details: The recombinant human CD274-Fc fusion is expressed as a 448 amino acid protein consisting of Phe19 - Arg238 region of CD274 (UniProt accession #Q9NZQ7) and a C-terminal Fc from human IgG1, which exists as a dimer under non-reducing conditions (see the gel image above). The purified protein is further biotinylated using standard procedure.

 

Source:  Human cells stably expressing PD-L1-Fc and growing in chemical-defined media with no animal components or antibiotics

 

Deduced Amino Acid Sequence:

FTVTVPKDLYVVEYGSNMTIECKFPVEKQLDLAALIVYWEMEDKNIIQFVHGEEDLKVQHSSYRQRARLLK

DQLSLGNAALQITDVKLQDAGVYRCMISYGGADYKRITVKVNAPYNKINQRILVVDPVTSEHELTCQAEGY

PKAEVIWTSSDHQVLSGKTTTTNSKREEKLFNVTSTLRINTTTNEIFYCTFRRLDPEENHTAELVIPELPL

AHPPNERSTGTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVE

VHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPS

REEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSC

SVMHEALHNHYTQKSLSLSPGK

M.W.: Calculated molecular mass 50.7 kDa; estimated by SDS-PAGE under reducing condition ~ 60 kDa suggesting that the protein is highly glycosylated 

 

Calculated PI: 6.27

 

Calculated Extinction Coefficients (M-1 cm-1, at 280nm):  61,935

 

Purity: >95% judged by SDS-PAGE under reducing condition (see the gel image above)

 

Formulation: Supplied at 0.5 mg/ml in sterile PBS pH7.4 (carrier free).

 

Endotoxin Level: <0.1 EU per 1 μg of purified recombinant protein determined by the LAL method

 

Biological Activity: Binds to its receptor PD-1-Fc (SKU# FCL0763) CD80-Fc (SKU# FCL0723) in functional ELISA (see Technical Data) or PD-1/CD80 molecule on cell surface with low affinity (K_D in μM range) and anti-PD-L1 monoclonal antibody (SKU# MAB0199) with high affinity (K_D < 1 nM).  Blocks PD-L1’s binding to its receptor and mediated signaling activity.

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

Restriction: This product is not transferable or re-sellable.  Customer obtain no right to transfer, assign, or sublicense its use rights, or to transfer, resell, package, or otherwise distribute the product, or to use the product for the benefit of any third party or for any commercial purpose.  Customer may only use the product in compliance with applicable local, state and federal laws, regulations and rules.  Customer may not directly or indirectly use the product or allow the transfer, transmission, export or re-export of all or any part of the product in violation of any export control law or regulation of the united states or any other relevant jurisdiction.  Your use of this product constitutes acceptance of the terms of this limited use agreement.  Please refer to our “terms & conditions” for details.  If you are not willing to accept the limitation of this agreement, G&P Biosciences will accept return of the product for a full/partial refund. 

 

  

Technical Data 1: Determination of Binding Affinity to Human PD-1 by ELISA

Recombinant PD-1-Fc protein (SKU#FCL0763) and Fc protein alone (SKU#FCL0029) was immobilized at 100 ng/well on a 96-well NuncMaxisorp plate for a functional ELISA with serially diluted biotinylated PD-L1-Fc protein (SKU#FCL0781B). Fc protein alone was used as a negative control to demonstrate the specificity of PD-1:PD-L1 binding under the experimental conditions.  The binding curve was fit and EC₅₀ (the concentration at 50% of max binding) was calculated for PD-1: PD-L1 binding to be 495 ± 74 ng/ml with a linear binding range between 50 and 2500 ng/ml.   

 

 

Technical Data 2: Determination of Binding Affinity to Human CD80 by ELISA

Recombinant CD80-Fc protein (SKU#FCL0723) and Fc protein alone (SKU#FCL0029) was immobilized at 100 ng/well on a 96-well NuncMaxisorp plate for a functional ELISA with serially diluted biotinylated PD-L1-Fc protein (SKU#FCL0781B). Fc protein alone was used as a negative control to demonstrate the specificity of CD80:PD-L1 binding under the experimental conditions.  The binding curve was fit and EC₅₀ (the concentration at 50% of max binding) was calculated for CD80 : PD-L1 binding to be 1967 ± 268 ng/ml with a linear binding range between 250 and 10,000 ng/ml.   

 

   

 

 FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

 

 

The product is shipped at 4°C. Upon receipt, centrifuge the product briefly before opening the vial. It is recommended to store small aliquots at the temperature below –20°C for long-term storage and the product is stable for 3 months. The undiluted protein can be stored at 4°C for no more than 2 weeks.

 

 

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

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

 

 

1. Nat. Med. 5:1365-1369, 1999

2. J. Exp. Med. 192:1027-1034, 2000

3. Nature Med. 8: 793-800, 2002

4. Nature Med. 9: 562-567, 2003

 

 

 

Product datasheet (pdf) can be downloaded here: FCL0781-PDS.pdf

 

 

 

Additional supporting documents, including COA and MSDS are available upon request.

 

 

 

FOR RESEARCH USE ONLY. NOT FOR DIAGNOSTIC OR THERAPEUTIC USE IN HUMAN.

Restriction: This product is not transferable or re-sellable.  Customer obtain no right to transfer, assign, or sublicense its use rights, or to transfer, resell, package, or otherwise distribute the product, or to use the product for the benefit of any third party or for any commercial purpose.  Customer may only use the product in compliance with applicable local, state and federal laws, regulations and rules.  Customer may not directly or indirectly use the product or allow the transfer, transmission, export or re-export of all or any part of the product in violation of any export control law or regulation of the united states or any other relevant jurisdiction.  Your use of this product constitutes acceptance of the terms of this limited use agreement.  Please refer to our “terms & conditions” for details.  If you are not willing to accept the limitation of this agreement, G&P Biosciences will accept return of the product for a full/partial refund.