| Description |
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The Immune Response Reporter Array is the first commercial reporter array
that allows you a comprehensive view of immune response signaling pathways.
Don't rely on guesswork to predict gene or drug function. Use the Cignal
Finder Immune Response Array to pinpoint the pathways regulated by the genes
or drugs your laboratory is studying.
Learn how the Immune Response Reporter Array works
The Cignal Finder Immune Response 10-Pathway Reporter Array is ideally
suited for discovering:
- siRNA/shRNA/miRNA phenotypes
- Biological responses to chemical compounds
- Mechanisms of action of proteins, peptides, and ligands
For more information about the
Cignal Finder 10-Pathway Reporter Arrays, please visit the Cignal Finder
Array home page
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| Pathway Listing and Performance Data |
| Tube/Column |
Pathway |
Transcription Factor |
| 1 |
NFκB |
NFκB |
| 2 |
PKC/Ca++ |
NFAT |
| 3 |
Type 1 Interferons |
STAT1/STAT2 |
| 4 |
Interferon Gamma |
STAT1/STAT1 |
| 5 |
MAPK/ERK |
Elk1/SRF |
| 6 |
MAPK/JNK |
AP1 |
| 7 |
TGFβ |
SMAD2/SMAD3/SMAD4 |
| 8 |
cAMP/PKA |
CREB |
| 9 |
C/EBP |
C/EBP |
| 10 |
Glucocorticoid Receptor |
GR |
| 11 |
Cignal negative control |
|
| 12 |
Cignal positive control |
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Learn more about the 10 pathways
View each reporter's transcriptional
response element sequence
Mammalian cells respond to infection through a complex immune response
involving both innate and adaptive immune systems. The 10
Cignal Reporter Assays included in the Cignal Finder Immune Response Reporter Array provide comprehensive examination of both components of the immune
response.
The NFκB transcription factor family controls B- and T-cell activation,
and inflammation. This pathway is frequently mutated in immune system
disorders and cancers. The cAMP/PKA signal transduction pathway leads to
activation of the CREB transcription factor, and works in concert with the NFκB pathways. The
cAMP/PKA pathway is a key convergence point for several
immunomodulators, including the PKC/Ca++, and TGFβ signals. The
C/EBP
transcription factors regulate a number of immune response genes. They have
been shown to bind to, and regulate the activities of, the NFκB and CREB
transcription factors. Corticosteroids are potent anti-inflammatory agents,
and are heavily used in the clinic and as a research tool. They mediate
their effects through the activation of Glucocorticoid Receptors. Many of
the growth factors, mitogens, cytokines, and chemokines that regulate immune
responses transmit their signals via the MAPK/ERK and MAPK/JNK signaling
pathways, impacting the activities of numerous immunoresponsive
transcription factors. These include NFκB, CREB, AP-1, ELK-1, and
STATs, all
of which can be monitored by the reporter assays included in this reporter
array. Activation of STATs can trigger the type 1 interferon and gamma
interferon responses. Activation of the type 1 interferon gene targets is
regulated by the Interferon-Stimulated Response Element (ISRE). Gamma
interferon induction results in the induction of genes controlled by the
Interferon-Gamma Activation Sequence (GAS).
Performance Data
Identify signaling pathway(s) regulated by recombinant protein
Tumor necrosis factor-alpha (TNF-α) is a pleiotropic inflammatory cytokine.
It is important to determine the key immunology signaling pathways modulated
by TNF-α. The Cignal Finder Immune Response 10-Pathway Reporter Array can
provide valuable information about the signaling pathways involved in the
biological response to TNF-α.
The Immune Response 10-pathway Reporter Array reveals that TNF-α
activates the NFκB and MAPK/JNK signaling pathways in HeLa cells.
HeLa cells were reverse transfected with the Cignal Immune Response
10-pathway Reporter Array. After 16 hours of transfection, medium was changed to
assay medium (Opti-MEM containing 0.5% of fetal bovine serum, 1% NEAA, 100 U/ml
Penicillin and 100 µg/ml Streptomycin). After 32 hours of transfection, cells
were treated with 5 ng/ml of TNF-α, or were left untreated. After 6 hours of
treatment, dual-luciferase assays were performed and results are expressed as
fold change. The fold change was calculated by dividing the normalized
luciferase activities of each pathway-focused reporter treated with TNF-α by the
normalized luciferase activity of the respective untreated pathway-focused
reporter. Experiments were done in quadruplicates, and the standard deviations
are indicated.
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| How It Works
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Each Reporter Array includes 10 Cignal Reporter Assays and two controls in
either tube or plate format. All reporter assays are based on dual-luciferase
technology. Each reporter consists of a mixture of a pathway-focused
transcription factor-responsive firefly luciferase construct and a
constitutively expressing Renilla luciferase construct.
Dual-luciferase results are calculated for each transfectant. The change in
the activity of each signaling pathway is determined by comparing the normalized
luciferase activities of the reporter in treated versus untreated transfectants.
The identically treated negative control serves as a specificity control. The
positive control serves as a control for transfection efficiency, by monitoring
GFP expression, as well as a positive control for both the firefly and Renilla
luciferase assays.
Simple Procedure:
- Transfect Cignal Reporter Assays and test nucleic acids into cells
- Treat with protein, peptide, or small molecule of interest
- Perform reporter quantitation using luciferase activity assays
Cignal Finder Array - Tube Format
The tube format of the Cignal Finder Arrays are delivered in 12-tube
strips, along with important negative and positive controls. The assays
are used right out of the box for the transfection or reverse
transfection of the reporter assays into your cell lines of interest.
Cignal Finder Array - Plate Format
The plate format of the Cignal Finder Arrays are delivered in a 96-well
cell culture plate. Each reporter and control assay is dried down in
each column of the plate (8 wells per assay).
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| Useful Links |
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| Related Products |
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| Brief Protocol: For Experienced
Users |
First time users, please refer to the complete
protocol in the Cignal Reporter Assays User Manual.
- Dilute transfection-ready reporter, negative control, and positive
control construct formulations.
- Dilute relevant test nucleic acids (siRNA, shRNA, miRNA, expression
vector).
- Prepare appropriate combinations of reporter constructs, controls,
and test nucleic acids.
- Transfect plasmid mixtures separately into replicate wells of your
cell line of interest using an optimized transfection procedure for
the cell line under study.
- If applicable, 16 to 24 hours post-transfection, treat the
transfected cells with test proteins, peptides, or compounds of
interest.
- Two (2) to three (3) days post-transfection, assay the activities of
the signaling pathways under study, utilizing the dual luciferase
assay.
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