PD1-CD28/NFAT Reporter Lentivirus

Background

PD-1 (PDCD1) is a co-inhibitory receptor on activated T cells that, upon engaging its ligands PD-L1 and PD-L2, dampens T cell receptor signaling and promotes peripheral tolerance and tumor immune evasion. CD28 is the principal co-stimulatory receptor that, by binding CD80 and CD86, amplifies T cell activation, proliferation, and survival. Engineered PD1-CD28 switch receptors fuse the PD-1 ectodomain to the CD28 signaling domain so that an inhibitory input is converted into a stimulatory one. Both signals converge on NFAT, a calcium-responsive transcription factor that drives effector gene expression downstream of the T cell receptor, making NFAT-based reporters a quantitative measure of T cell activation in checkpoint biology.

Product Description & Applications

The PD1-CD28/NFAT Reporter Lentivirus is a two-component immunotherapy reporter system. A receptor construct uses an EF1a promoter to constitutively co-express human PD1 and CD28, separated by a P2A linker, together with a fluorescent reporter or drug selection marker for stable cell line selection. A separate cassette drives a reporter through tandem NFAT response elements coupled to a minimal promoter and upstream enhancer, yielding a fluorescent (GFP or RFP) and/or secreted Gaussia luciferase readout of T cell activity.

Sequential transduction generates dual-stable effector lines for studying PD-1 pathway modulation, checkpoint-blockade agents, and switch-receptor engineering. Supplied as high-titer, VSV-G-pseudotyped particles purified by PEG precipitation and sucrose gradient centrifugation, suitable for primary and thawed cells.

About This Product

This 2-vial immunotherapy reporter system consists of a Vial 1 Receptor Lentivirus encoding human PD1-P2A-CD28 under a constitutive promoter with antibiotic selection, and a Vial 2 Reporter Lentivirus encoding tandem NFAT (or NF-κB) response elements driving a dual reporter (GFP, GFP-P2A-GLuc, GLuc, RFP, RFP-P2A-GLuc). Sequential transduction and selection generates a dual-stable effector cell line that responds quantitatively to receptor stimulation with a ratiometric fluorescent + bioluminescent readout.

Secreted Gaussia luciferase (where included) accumulates in conditioned media, enabling kinetic sampling without cell lysis. The combined fluorescent and luminescent outputs allow parallel microscopy-based visualization and plate-reader luminometry from the same cell population — providing assay redundancy and flexibility for potency testing formats compliant with regulatory expectations for cell-based functional assays.