Cellsensor Cellular Pathway Assays

CellSensor™ c-Fos-bla ME-180 Cell Line Thermo Scientific™

The CellSensor® c-fos-bla ME-180 cell line contains a beta-lactamase reporter gene under control of the c-fos response element stably integrated into ME-180 cells. To construct this cell line, the c-fos-bla construct was transduced into ME-180 cells by lentivirus. Flow cytometry was used to isolate cells responsive to epidermal growth factor (EGF). This cell line is validated for DMSO tolerance, cell number, stimulation time, substrate loading time, as well as Z' and EC50 concentrations for EGF. The CellSensor® c-fos-bla ME-180 cell line is responsive to EGF, HGF, IL-6, OSM, TNFα, IL-1-alpha; and PMA/Thaps, and can be used to probe the JNK/P38/MAPK and JAK/STAT signaling pathways. This cell line can be adapted for high-throughput screening of agonist or antagonist compound libraries. Candidate drugs can also be tested for dose response against this cell line. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ c-Fos-bla HEK293T Cell Line Thermo Scientific™

The c-fos-bla HEK 293T cell line contains a beta-lactamase reporter gene under control of the c-fos promoter stably integrated into HEK 293T cells. This cell line can be used to detect agonists/antagonists of the c-fos pathway. The c-fos-bla HEK 293T cells have been shown to be responsive to phorbol 12-myristate 13-acetate (PMA) stimulation. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ GAS-bla ME-180 Cell Line Thermo Scientific™

The CellSensor™ GAS-bla ME-180 cell line contains a beta-lactamase reporter gene under control of the Gamma Activated Sequence (GAS) response element stably integrated into ME-180 cells. To construct this cell line, the GAS-bla construct was transduced into ME-180 cells by lentivirus. Flow cytometry was used to isolate cells responsive to Interferon Gamma (IFN-γ). This cell line is validated for DMSO tolerance, cell number, stimulation time, substrate loading time, as well as Z'and EC50 concentrations for IFN-&gamma. The CellSensor™ GAS-bla ME-180 cell line is responsive to IFN-γ and can be used to probe the JAK/STAT signaling pathway. This cell line can be adapted for high-throughput screening of agonist or antagonist compound libraries. Candidate drugs can be tested for dose response against this cell line. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ TNFa-bla THP-1 Cell Line Thermo Scientific™

Tumor necrosis factor-alpha (TNFα) cytokine cell signaling is a critical pathway involved in immune and inflammatory responses. In addition, TNFα has been shown to inhibit growth and induce differentiation in tumor and non-tumor cells. The CellSensor™ TNFα-bla THP-1 cell line is a suspension cell line derived from human acute monocytic leukemia cells. The TNFα-bla THP-1 cell line is used to measure the TNFα cytokine signaling pathway via transcriptional regulation of the beta-lactamase gene. This cell line was constructed using GenomeScreen™ technology utilizing a promoterless beta-lactamase reporter gene randomly integrated into the genome of the THP-1 cell line. Sequential rounds of sorting were performed to isolate cell clones containing tagged genes induced by TNFα. These cells have been shown to be responsive to TNFa agonist. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ AP-1-bla A375 Cell Line Thermo Scientific™

The CellSensor® AP1-bla A375 cell line contains a beta-lactamase reporter gene under control of the AP1 response element stably integrated into A375 cells. A375 cells are human melanoma cancer cells that contain the endogenous B-Raf mutation V600E resulting in constitutive B-Raf kinase activity. The CellSensor® AP1-bla A375 cell line is a clonal population isolated by flow cytometry based on constitutive expression of beta-lactamase. This cell line has been validated with various small-molecule Raf inhibitors as well as B-Raf Stealth RNAi™ siRNA. This cell line has also been tested for assay performance under variable conditions, including DMSO concentration, cell number, compound incubation time, and substrate loading time and validated for Zfi-factor and IC50 concentrations of Raf1 Inhibitor I. The RAF gene family (RAF1, A-RAF, and B-RAF) encodes closely related serine/threonine protein kinases that are important effectors of Ras activation. Raf1 and A-Raf are rarely mutated, whereas mutations in B-Raf are common in human cancers, especially melanoma. B-Raf is mutated in about 70% of human melanomas, 35-70% of papillary thyroid carcinomas, and less commonly in lung and colorectal carcinomas. Mutations are mostly in the B-Raf kinase domain and, in melanomas, the vast majority are V600E missense mutations leading to activation of B-Raf kinase. The constitutive activity of B-Raf V600E can directly lead to the activation of the Mek/MAPK signaling pathway. Therefore, inhibition of B-Raf/Mek/MAPK signaling could be a potential way of treating melanomas and other tumors with mutant B-Raf.

CellSensor™ NFAT HEK 293T DA Assay Kit Thermo Scientific™

The CellSensor® NFAT HEK 293T DA (Division Arrested) cells contain a beta-lactamase reporter gene under the control of the Nuclear Factor of Activated T Cells (NFAT) response element stably integrated into HEK 293T cells. Fluorescence activated cell sorting (FACS) was used to isolate clones responsive to stimulation of the NFAT pathway by PMA (Phorbol 12-Myristate 13-Acetate) and Thapsagargin (Thaps). The cell line was validated for DMSO tolerance, cell number, stimulation time, substrate loading conditions, and EC50 concentration of Thaps at a fixed concentration of PMA. DA cells are irreversibly division arrested using a low-dose treatment of Mitomycin C, and have no apparent toxicity or change in cellular signal transduction.

This product can serve as a negative control in screening assays performed with specific G-protein coupled receptor (GPCR) DA cells that were made from this same background or these cells can be used in other assays where a sensitive readout to intracellular changes in calcium are needed.

CellSensor™ irf1-bla Ba/F3 Cell Line Thermo Scientific™

Jak/Stat signaling pathways play essential roles in the cellular responses to distinct cytokines. One of the Jak/Stat pathways, Jak2/Stat5, mediates cell proliferation in response to Interleukin - 3 (IL - 3), prolactin, erythropoietin (Epo), and granulocyte - macrophage colony stimulating factor (GM - CSF). JAK2 gene knockout causes embryonic lethality due to defective erythropoiesis, suggesting that the Jak2/Stat5 pathway plays an important role in red blood cell formation. The recent discovery of an activating mutation in JAK2 (V617F) present in a high percentage of myeloproliferative disease (MPD) patients suggests that the Jak2/Stat5 pathway may be a potential therapeutic target for certain forms of MPD. The activated Stat5 transcription factor recognizes and binds to a specific palindromic DNA sequence found in the promoter region of β - casein, interferon regulatory factor - 1 (irf - 1), and a number of other genes. The CellSensor® irf1 - bla BA/F3 Cell Line contains a beta - lactamase reporter gene under control of the irf - 1 response element stably integrated into BA/F3 cells. This cell line is validated for EC50 and Z' - factor under optimized conditions using mouse IL - 3 (mIL - 3). This cell line has also been tested under variable experimental conditions, including DMSO concentration, cell number, stimulation time, and substrate loading time. Responsiveness to Jak Inhibitor 1, a small molecule inhibitor, was also tested. CellSensor® irf1 - bla BA/F3 cells were treated with mIL - 3 in triplicate over the indicated concentration range in a 384 - well format. Cells were incubated for 5 hrs with mIL - 3 agonist in 0.5% DMSO and then combined with LiveBLAzer™ - FRET B/G Substrate (CCF4 - AM) for 2.5 hrs. Fluorescence emission values at 460 nm and 530 nm were obtained using a standard fluorescence plate reader. The Response Ratios for each replicate were plotted against the indicated concentrations of mIL - 3. CellSensor ® irf1 - bla BA/F3 cells were treated with Jak Inhibitor 1 for 30 min over the indicated concentration range in a 384 - well format. Cells were then incubated with mIL - 3 agonist in 0.5% DMSO for 5 hrs and then combined with LiveBLAzer™ - FRET B/G Substrate (CCF4 - AM) for 2.5 hrs. Fluorescence emission values at 460 nm and 530 nm were obtained using a standard fluorescence plate reader. The emission ratios were plotted against the indicated concentrations of Jak Inhibitor 1. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ T-REx™ NICD CSL-bla HeLa Cell Line Thermo Scientific™

The CellSensor® T-REx™ NICD CSL-bla HeLa cell line was engineered by lentiviral transduction of HeLa cervical cancer cells with a notchresponse element driving beta-lactamase reporter gene expression (CSL-bla) along with tetracycline repressor and tetracycline (or the tetracycline analog, doxycycline)-inducible NICD (notch intracellular domain) constructs. This cell line is a clonal population isolated by flow cytometry. Addition of doxycycline to these cells allows for regulated NICD transcription factor expression and subsequent beta-lactamase expression. This cell line has been tested for robust assay performance by assessing a variety of parameters, including cell plating number, DMSO tolerance, stimulation time, and substrate loading time. Assay validation was performed using serial dilutions of doxycycline. Additional testing data using RNAi and alternate stimuli are also available.

CellSensor™ NFkB-bla RAW 264.7 Cell Line Thermo Scientific™

The CellSensor® NFκB-bla RAW264.7 mouse macrophage cell line contains a beta-lactamase reporter gene under control of a stably integrated NFκB response element. This cell line is a clonal population isolated in response to lipopolysaccharide by flow cytometry. This cell line has been tested for assay performance under variable conditions, including DMSO concentration, cell number, stimulation time, and substrate loading time, and has been validated for Zfi-factor and EC50 concentrations of the TLR7 ligand imiquimod. Additional data using alternate stimuli are shown. The molecules responsible for coordinating the immune systems recognition of pathogens are Toll-like receptors (TLRs), of which there are currently 10 identified members in humans. TLRs are pattern recognition receptors (PRRs), which bind to pathogen-associated molecular patterns (PAMPs), small molecular sequences consistently found on pathogens. This binding activates the transcription of immune system genes and regulators such as cytokines, chemokines, and costimulatory molecules, thereby promoting the initial immune response of macrophages and neutrophils. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ IL1-bla ECV304 Cell Line Thermo Scientific™

The CellSensor® IL-1-bla ECV304 cell line is an adherent human urinary bladder cell line used to measure the (IL-1) signaling pathway via transcriptional regulation of the beta-lactamase gene. IL-1 cytokine cell signaling is a critical pathway involved in immune and inflammatory responses. This cell line was constructed using GenomeScreen™ technology utilizing a promoterless beta-lactamase reporter gene randomly integrated into the genome of the IL-1-bla ECV304 cell line. Sequential rounds of sorting were performed to isolate cell clones containing tagged genes induced by IL-1. A dose response assay was run with this cell line using IL-1 as the agonist. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ SBE-bla A375 Cell Line Thermo Scientific™

Bone morphogenetic proteins (BMPs), members of the transforming growth factor beta (TGF-β) superfamily, play important roles in the development of the heart, central nervous system and cartilage. Disruption of BMP signaling affects the body plan of the developing embryo. BMPs propagate their signal by activating Activin receptor-like kinase (ALK), which in turn mediates the phosphorylation of R-Smads. Phosphorylated R-Smad associates with Smad4 and then translocates to the nucleus regulating gene expression. A375 cells are human malignant melanoma cells containing endogenous Smad signaling pathway. SBE-bla A375 cell line was engineered to express beta-lactamase under the control of Smad binding element. This cell line has been validated for Z' and EC50 under optimized conditions using BMP-4. This cell line has also responded to Nodal and was tested for assay performance under variable conditions, including DMSO concentration, cell number, stimulation time, substrate loading time. Additional information using various small molecule inhibitors and StealthTM RNAi is also provided.

CellSensor™ AP-1-bla ME-180 Cell Line Invitrogen™

The CellSensor® AP-1-bla ME-180 Cell Line contains a beta-lactamase reporter gene under control of the AP-1 response element stably integrated into ME-180 cells. The cell line was created through FACS sorting of cells responsive to stimulation of the AP-1 pathway with Epidermal Growth Factor (EGF). This cell line was validated for DMSO tolerance, stimulant incubation time, substrate loading conditions, Z' and EC50 concentration of Epidermal Growth Factor. The AP-1-bla ME-180 Cell Line responds to agonist treatment as expected from the literature and can be adapted for high-throughput screening for agonists or antagonists of the AP-1 pathway with compound libraries. Candidate drugs can also be tested for dose response against this cell line. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ CRE-bla CHO K1 Cell Line Thermo Scientific™

The CellSensor® CRE-bla CHO-K1 cell line contains a beta-lactamase reporter gene under control of a cAMP response element (CRE) stably integrated into CHO-K1 cells. This cell line can be used as a parental cell line to build specific G-protein coupled receptor (GPCR) assays after transfection of additional genes of interest or to detect changes in intracellular cAMP levels. CRE-bla CHO-K1 cells have been shown to be responsive to forskolin stimulation. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ irf1-bla TF-1 Cell Line Thermo Scientific™

Jak/Stat signaling pathways play essential roles in cellular responses to distinct cytokines. One of the Jak/Stat pathways, Jak2/Stat5, mediates cell proliferation in response to Interleukin - 3 (IL - 3), prolactin, erythropoietin (Epo), and granulocyte - macrophage colony stimulating factor (GM - CSF). JAK2 gene knockout causes embryonic lethality due to defective erythropoiesis, suggesting that the Jak2/Stat5 pathway plays an important role in red blood cell formation. The recent discovery of an activating mutation in JAK2 (V617F) present in a high percentage of myeloproliferative disease (MPD) patients suggests that the Jak2/Stat5 pathway may be a potential therapeutic target for certain forms of MPD. The activated Stat5 transcription factor recognizes and binds to a specific palindromic DNA sequence found in the promoter region of β - casein, interferon regulatory factor - 1 (irf - 1), and a number of other genes. The CellSensor® irf1 - bla TF1 Cell Line contains a beta - lactamase reporter gene under control of the irf - 1 response element stably integrated into TF1 cells. TF1 cells are a human erythroleukemia cell line that is growth - dependent on GM - CSF and have an intact GM - CSF - JAK2 - STAT5 pathway. This cell line is validated for EC50 and Z’ - factor using GM - CSF. This cell line has also been tested under variable experimental conditions, including DMSO concentration, cell number, stimulation time, and substrate loading time. Responsiveness to EPO and Jak Inhibitor 1 was also tested (Figures 2 and 3). CellSensor® irf1 - bla TF1 cells were stimulated in triplicate with GM - CSF over the indicated concentration range in a 384 - well format. Cells were incubated for 5 hrs with agonist and 0.5% DMSO and then combined with LiveBLAzer™ - FRET B/G Substrate (CCF4 - AM) for 2.5 hrs. Fluorescence emission values at 460 nm and 530 nm were obtained using a standard fluorescence plate reader. The 460/530 ratios were plotted for each replicate against the indicated concentrations of GM - CSF. CellSensor® irf1 - bla TF1 cells were stimulated with Epo over the indicated concentration range in a 384 - well format. Cells were incubated for 5 hrs with agonist and 0.5% DMSO and then combined with LiveBLAzer™ - FRET B/G Substrate (CCF4 - AM) for 2.5 hrs. Fluorescence emission values at 460 nm and 530 nm were obtained using a standard fluorescence plate reader. The Response Ratios were plotted against the indicated concentrations of Epo. CellSensor® irf1 - bla TF1 cells were treated with Jak Inhibitor 1 over the indicated concentration range in a 384 - well format. Cells were then stimulated with GM - CSF or EPO for 5 hrs in 0.5% DMSO and then combined with LiveBLAzer™ - FRET B/G Substrate (CCF4 - AM) for 2.5 hrs. Fluorescence emission values at 460 nm and 530 nm were obtained using a standard fluorescence plate reader. These values were converted to percent inhibition relative to a set of controls (unstimulated cells and EC80 of GM - CSF or EPO treated cells) and plotted against the indicated concentrations of Jak Inhibitor 1. Academic and non-profit customers, please inquire for special pricing.

CellSensor™ Gli-bla 22Rv.1 Cell Line Thermo Scientific™

The morphogenic signal Shh provides in the developing CNS induces proliferation of neuronal precursor cells in the developing cerebellum and other tissues. Proliferative signaling by Shh is involved in the development of cancer, including specific brain and skin cancers such as basal cell carcinomas. Signaling takes place through a Patched (PTC-1)⁄Smoothened (SMO) Receptor complex. The activation of Patched by Shh releases the inhibition of Patched on Smoothened leading to Sonic Hedgehog pathway activation of the Gli transcription factor to induce downstream gene expression. The CellSensor® Gli-bla 22Rv1 cell line contains a beta-lactamase reporter gene under control of the Gli response element stably integrated into 22Rv1 cells. This cell line is a clonal population isolated by flow cytometry. The clone was selected based on inhibition of the pathway with KAAD-Cyclopamine. This cell line has also been tested for assay performance under variable conditions, including DMSO concentration, cell number, compound incubation time, and substrate loading time and validated for Z’ under maximal inhibition of β-lactamase activity with Clavulanic Acid.
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