NFAT eGFP Reporter Lentivirus

Catalog #

79922

$910 *

Size: 500 µl x 2

Qty

Bulk/Custom

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*US Pricing only. For international pricing, please contact your local distributor.

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Description

NFAT eGFP Reporter Lentivirus are replication incompetent, HIV-based, VSV-G pseudotyped lentiviral particles that are ready to transduce almost all types of mammalian cells, including primary and non-dividing cells. The particles contain an enhanced GFP (green fluorescent protein) reporter driven by the NFAT (Nuclear Factor of Activated T Cells) response element located upstream of the minimal TATA promoter. The lentiviruses also transduce a puromycin selection gene (Figure 1). After transduction, activation of the NFAT signaling pathway in target cells can be monitored by assessing eGFP expression.

Figure 1. Schematic of the lenti-vector used to generate the NFAT eGFP reporter lentivirus

Figure 1. Schematic of the lenti-vector used to generate the NFAT eGFP Reporter Lentivirus.

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Product Data Gallery

NFAT eGFP reporter activity stimulated by Anti-CD3 Agonist Antibody in Jurkat cells transduced with NFAT eGFP Reporter Lentivirus

Product Info

Storage and Usage

Citations

Supplied As

Two vials (500 µl x 2) of NFAT eGFP Reporter Lentivirus at ≥10⁷ TU/ml. The titer will vary with each lot; the exact value is provided with each shipment.

Materials Required But Not Supplied

Jurkat Cells (ATCC #TIB-152)
Thaw Medium 2 (BPS Bioscience #60184)
Lenti-Fuse™ Polybrene Viral Transduction Enhancer (BPS Bioscience #78939)
Anti-CD3 Agonist Antibody (BPS Bioscience #71274)
ONE-Step™ Luciferase Assay System (BPS Bioscience #60690)
96-well white clear-bottom assay plate (Corning #3610)

Formulation

The lentiviruses were produced from HEK293T cells. Supplied in medium containing 90% DMEM + 10% FBS. Virus particles can be packaged in custom formulations and produced at higher titers by special request, for an additional fee.

Background

The NFAT (Nuclear Factor of Activated T Cells) family of transcription factors plays an important role in mediating immune responses. T cell activation through the T cell synapse results in calcium influx, which is required to facilitate actin organization and interactions at the synapse. Increased intracellular calcium levels activate the calcium-sensitive phosphatase calcineurin, which rapidly dephosphorylates the serine-rich region (SRR) and SP-repeats in NFAT proteins. This results in a conformational change that exposes a nuclear localization signal (NLS), promoting NFAT nuclear translocation and inducing gene expression, including various cytokines (IL-2, IL-3, IL-4, and TNFα). Members of the NFAT family have been found in many tissue types, including the heart, skeletal muscle, and brain. These transcription factors are known to be highly involved in the pathological processes of inflammation and cancer. The use of eGFP (enhanced green fluorescent protein) allows for easy assay readouts.

References

Clipstone N.A., et al., 1992 Nature 357(6380): 695-697.
Lyakh L., et al., 1997 Mol Cell Biol. 17(5): 2475-2484.

Storage/Stability

Lentiviruses are shipped with dry ice. For long-term storage, it is recommended to store the lentiviruses at -80°C for up to 12 months from date of receipt. Avoid repeated freeze/thaw cycles. Titers can drop significantly with each freeze/thaw cycle.

Applications

Screen for activators or inhibitors of NFAT signaling pathway in transduced target cells.
Generate NFAT eGFP reporter cell pools or stable cell lines following puromycin selection.

Shipping Temperature

-80°C

Notes

To generate a NFAT expressing stable cell line, remove the growth medium 48 hours after transduction and replace it with fresh growth medium containing the appropriate amount of puromycin (as pre-determined from a killing curve, https://bpsbioscience.com/cell-line-faq), for antibiotic selection of transduced cells, followed by clonal selection.

Troubleshooting Guide
Visit bpsbioscience.com/lentivirus-faq for detailed troubleshooting instructions. For all further questions, please email [email protected].

Biosafety
The lentiviruses are produced with the SIN (self-inactivation) lentivector which ensures self-inactivation of the lentiviral construct after transduction and integration into the genomic DNA of the target cells. None of the HIV genes (gag, pol, rev) will be expressed in the transduced cells, as they are expressed from packaging plasmids lacking the packing signal. Although the pseudotyped lentiviruses are replication-incompetent, they require the use of a Biosafety Level 2 facility. BPS recommends following all local federal, state, and institutional regulations and using all appropriate safety precautions.

Warnings

Avoid freeze/thaw cycles

Scientific Category

Cell Signaling Pathway

Regulatory Status

Research Use Only

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NFAT eGFP Reporter Lentivirus

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