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CD16A/NFAT Reporter Lentivirus (ADCC Assay)
$895.00 – $1,695.00
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SIRPA/NFκB Reporter Lentivirus
$895.00
CD32A/NFAT Reporter Lentivirus (ADCP Assay)
$895.00 – $1,695.00
The CD32/NFAT Immunotherapy Reporter Lentivirus is designed to evaluate the potency of antibody-dependent cellular phagocytosis (ADCP), a crucial Fc effector function mediated by CD32 on the surface of macrophages. When the CD32 receptor binds to the Fc region of a specific antibody targeting cells, it triggers internalization and degradation of the target cells through phagosome acidification. Detecting phagosome activity in ADCP is challenging in vitro. This cell-based reporter measures the association between target cells and effector macrophages, thereby assessing the ADCP effect.
Similar to ADCC assays, the exogenous expression of human CD32 with high affinity (H131 allotype) or low affinity (R131 allotype) on human T-lymphocyte lines, such as the Jurkat cell line, can measure NFAT activation and drive the expression of dual reporters: secreted Gaussia luciferase (GLuc) and GFP. The construct constitutively expresses the human CD32A high-affinity (H131) allotype and is intended to be used along with our companion product which expresses the low-affinity CD32A allotype R131, serving as a control. Transduce your desired cells with the all-in-one high-titer purified lentivirus to establish a stable, more effective cell line for ADCP assays.
This product is supplied as high-titer lentiviral particles (third generation), which have been purified by PEG precipitation and sucrose gradient centrifugation. The lentiviral particles are VSV-G pseudotyped and are ideal for difficult-to-transfect cells, including primary or thawed cells.
Have questions about this product? Need a stable cell line? Contact Us
Available Options:
Specifications
Details
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| TRV-0001 | |
| High-titer Lentiviral Particles, third generation, VSV-G pseudotyped | |
| Serum-free RPMI-1640, frozen solution | |
| One vial | |
| 380 μl/vial | |
| 1-2×10^6 TU/vial, depending on the variant | |
| 3×10^8 VP/ml | |
| -80oC | |
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| Promoter | EF1α |
| Human CD32A, H131 (C-terminal Flag) | |
| NM_001136219 | |
| Puromycin (Puro) or Blasticidin (BSD) | |
| Tandem repeats of response elements coupled to minimal TATA promoter and upstream proprietary enhancer | |
| NFAT response elements | |
| GFP or RFP plus secreted Gaussia Luc (GLuc), separated by P2A | |
| Antibody dependent cell-mediated phagocytosis (ADCP) assay. The product can be used to generate stable cell lines that read out CD32A(H131) signaling through downstream activity of NFAT. The fluorescent and luminescent dual reporter provides flexibility in the readout approach. GLuc is secreted, allowing the user to detect it in various biological media. Generally, the product is useful for analyzing cell activity as a function of CD32A stimulation/inhibition. |
*Transduction units (TU), based on the number of drug resistant or fluorescent HEK293FT cells after transduction with the virus.
Vector Layout
Product Data
Figure 1. Jurkat cells (5×10^5) were transduced with 100 μl of the CD32A(H131)-BSD/NFAT-GFP-GLuc lentiviral particles (SKU: TRV-0006-6S) in a 12-well plate with 6 μg/ml polybrene, followed by centrifugation for 2 hours. The cells were then incubated for an additional 2-4 hours. The entire culture was transferred to a T25 flask and cultured in 10 ml RPMI1640 medium for 3 days. The medium was then changed, and 10 μg/ml blasticidin was added to select for drug-resistant ADCP-Jurkat cells. These cells were co-cultured with target Raji cells in the presence of either a CD20 antibody (BioXCell SKU: SIM0008) or an isotype control antibody for 16 hours before imaging and flow cytometry analysis. Figure 1 on the left shows the fluorescence microscopy images obtained. The panels on the right side labeled “phase” show the cells with added light to allow visualization of the non-fluorescent cells. In the product thumbnail at the top of the page, flow cytometry profiles are depicted for the same experiment, showing the versatility of the fluorescent readout for the study of ADCP.
Figure 2. Same experiment as shown in Figure 1 with Jurkat-ADCP and Raji cells. GLuc expression was detected in the cell culture medium sampled at the same time the fluorescence images of the cells were taken. Twenty ul of the cell culture medium was combined with the assay reagents from the Promega Stop&Glow assay kit (Promega SKU: E1910) according to the manufacturer’s instructions and bioluminescence was detected on a SpectraMax iD3 (integration time 1 sec).
Figure 2. Drug-resistant ADCP-Jurkat cells were co-cultured with target Raji cells in the presence of 100 ng/ml of anti-CD20 antibody (BioXCell SKU: SIM0008) or an isotype control antibody for 16 hours before flow cytometry analysis.
Advantages
- Versatile Readouts Luminescent and fluorescent dual readout provide unprecedented flexibility.
- Open Doors to Discovery Monitor signal pathway activity in vitro or in vivo with secreted GLuc.
- Innovative Technology Utilizes our novel reporter technology with upstream enhancer to boost signal intensity.
- License-free & Affordable Generate your own cell lines with ease, freeing you from constraints of expensive fixed commercial reporter cell lines. The EF1α promoter-driven receptor/ligand provides long-term stable expression in cell lines.
- Unmatched customization Various reporters and selection markers to choose from, perfectly tailored to your specific research needs.
Custom Orders
If you require a modification to one of our products, such as a change in reporter or other vector component, please contact us. Examples of potential customization options are shown in the table below and can be custom-made upon request. Please note that not all options may be feasible due to size limitations and this will depend on the specific product. We also welcome requests for custom products like the one shown on this page.
Additional Custom Service Options
- Send us your cells and we can establish a stable reporter cell line for you using this product. Learn more.
- ORF cDNA plasmids featured in the product figures are available upon request.
- Ultra-high concentration virus can be provided upon request in your choice of medium and volume (i.e. for in vivo applications).
Lentiviral Transduction Protocol
MSDS
Frequently Asked Questions
Frequently Asked Questions
Can I purchase the plasmid for this product?
TF reporter products such as this one are not available in plasmid format. They are only available as high-titer lentiviral particles. Other gene products on our site, such as ORF cDNA and shRNA products are available in either format.