TCR Activator / PD-L1 Mammalian Expression Kit

Catalog #: 60610
Size: 500 reactions
$575
Species: Human
Background:
The binding of Programmed Cell Death Protein 1 (PD-1), a receptor expressed on activated T cells, to its ligands, PD-L1 and PD-L2, negatively regulates immune responses. The PD-1 ligands are found on most cancers, and PD-1:PD-L1/2 interaction inhibits T cell activity and allows cancer cells to escape immune surveillance. The PD-1:PD-L1/2 pathway is also involved in regulating autoimmune responses, making these proteins promising therapeutic targets for a number of cancers, as well as multiple sclerosis, arthritis, lupus, and type I diabetes.
Description:

The recombinant expression vectors are designed to express human engineered T cell receptor (TCR) activator and human PD-L1 (GenBank Accession #NM_014143) in mammalian cells. The transfected cells can be used in conjunction with PD-1/NFAT Reporter/Jurkat T cells (BPS #60535) to study the interactions of PD-1 with PD-L1 ligand in a cellular context and screen for modulators of this signaling pathway.

In this assay, PD-1/NFAT Reporter/Jurkat T cells are used as effector cells; HEK293 cells overexpressing PD-L1 (or PD-L2) and an engineered T cell receptor (TCR) activator are used as target cells. When these two cells are co-cultivated, TCR complexes on effector cells are activated by TCR activator on target cells, resulting in expression of the NFAT luciferase reporter. However, PD1 and PD-L1 (or PD-L2) ligation prevents TCR activation and suppresses the NFAT-responsive luciferase activity. This inhibition can be specifically reversed by anti-PD1 or anti-PD-L1 antibodies. PD1/PD-L1 neutralizing antibodies block PD1:PD-L1 interaction and promote T cell activation, resulting in reactivation of the NFAT responsive luciferase reporter.

Materials Required But Not Provided:

• HEK293 cell and its growth medium or other cell lines 
• Transfection reagent for mammalian cell line [BPS uses Lipofectamine™ 2000 (life technologies #11668027). However, other transfection reagents work equally well.]
PD-1/NFAT Reporter Jurkat T cells (BPS Cat. #60535)
• Opti-MEM I Reduced Serum Medium (life technologies #31985-062)
• Assay medium: RPMI1640 + 10% FBS + 1% Penicillin/Streptomycin 
Anti-PD-1 neutralizing antibody (BPS Cat. #71120) 
Anti-PD-L1 neutralizing antibody (BPS Cat. #71213) 
• 96-well tissue culture-treated white clear-bottom assay plate 
One-Step luciferase assay system (BPS Cat. # 60690) or other luciferase reagents for measuring firefly luciferase activity • Luminometer

Synonym(s):
Programmed Cell Death 1 Ligand 1, PDL1, PD-L1, B7 homolog 1, B7H1, B7-H1, CD274, PDCD1L1
Format:

Components

Instructions for use:
See assay kit protocol for detailed instructions.
Storage / Stability:
Stable at least 6 months from date of receipt, when stored as directed.
Application(s):
• Screen for activators or inhibitors of PD-1 signaling in a cellular context • Characterize the biological activity of PD-1 and its interactions with ligands
Notes:

Materials Required but Not Supplied
• HEK293 cell and its growth medium or other cell lines
• Transfection reagent for mammalian cell line [We use Lipofectamine™ 2000 (life technologies #11668027). However, other transfection reagents work equally well.] • PD-1/NFAT Reporter Jurkat T cells (BPS #60535) • Opti-MEM I Reduced Serum Medium (life technologies #31985-062)
• Assay medium: RPMI1640 + 10% FBS + 1% Penicillin/Streptomycin
• Anti-PD-1 neutralizing antibody: BPS bioscience #71120
• Anti-PD-L1 neutralizing antibody: BPS bioscience #71213
• 96-well tissue culture-treated white clear-bottom assay plate
• One-Step luciferase assay system (BPS bioscience # 60690) or other luciferase reagents for measuring firefly luciferase activity • Luminometer

Warning(s):
Avoid freeze/thaw cycles.
Scientific Category:
Immunotherapy
Product Type:
Assay Kit
Data shown is lot-specific. Contact us for specific information on other lots.

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Product Datasheets

60610.pdf