Cell signaling
Cellular signaling allows cells to respond to their environment and communicate with other cells. Proteins located on the cell surface can receive signals from the surroundings and transmit information into the cell via a series of protein interactions and biochemical reactions that comprise a signaling pathway. Multicellular organisms rely upon an extensive array of signaling pathways to coordinate the proper growth, regulation, and functioning of cells and tissues. If signaling between or within cells is dysregulated, inappropriate cellular responses may lead to cancer and other diseases.
Cell signaling is the collective name for processes, which cells receive signals, interpret, and respond accordingly. These signals range from physical cues, such as temperature, light, or electricity, to chemical signals, such as small molecules binding to the cell surface. With the ability to process these signals, cells can interact with each other and the environment. In addition, cell signaling constitutes the foundation of development, immune response, and homeostasis in all organisms. This is how our nerve cells communicate with our muscles to facilitate movement or how our body initiates an immune response upon recognizing pathogens.
Cellular responses
Many tools have been developed to measure cellular responses occurring through a wide range of signaling pathways. G protein-coupled receptor (GPCR) signaling can be studied using assays ranging from calcium flux, which can be monitored using fluorescence dyes, to changes in downstream effector molecules assessed by TR-FRET. Fluorescent molecules have been harnessed to reveal changes in the activity of kinases that play key roles in signaling pathways. Luciferase reporter gene assays utilizing luminescent reactions have become a versatile tool for studying gene regulation and cell signaling in both basic research and drug discovery.
The detection of cellular signals and responses can help us elucidate the role of cell-to-cell interactions in diseases. Drug discovery can also greatly benefit from cell signaling assays since many drugs inhibit aberrant ligand-protein interactions during signal transduction.
Cellular signaling applications and assays
Below, we present a variety of cell signaling applications that can be run using microplate readers with fluorescence, TR-FRET, fluorescence polarization, and luminescence detection modes.
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Calcium and IP-One assays (GPCR, Gq coupled)
Cell signaling via G protein-coupled receptors (GPCRs) can be assessed by monitoring the downstream effectors calcium or cyclic AMP (cAMP). Calcium flux in response to Gq protein-coupled receptor activation is commonly monitored in live cells in real time using calcium-sensitive dyes on a fluorescence plate reader. Another arm of this pathway is monitored using a TR-FRET assay for IP1.
Here are a few application notes you may find of interest:
- Monitoring Gq protein-coupled receptor activation on the SpectraMax i3x reader with injector module
- Optimization of a muscarinic M3-receptor assay using frozen CHO cells on the FlexStation 3 reader
- Calcium signaling with FLIPR Calcium 6 and 6-QF Assay Kits on the FlexStation 3 reader
- HTRF IP-One Gq assay on SpectraMax readers
Calcium and IP-One assays (GPCR, Gq coupled)
Cell signaling via G protein-coupled receptors (GPCRs) can be assessed by monitoring the downstream effectors calcium or cyclic AMP (cAMP). Calcium flux in response to Gq protein-coupled receptor activation is commonly monitored in live cells in real time using calcium-sensitive dyes on a fluorescence plate reader. Another arm of this pathway is monitored using a TR-FRET assay for IP1.
Here are a few application notes you may find of interest:
- Monitoring Gq protein-coupled receptor activation on the SpectraMax i3x reader with injector module
- Optimization of a muscarinic M3-receptor assay using frozen CHO cells on the FlexStation 3 reader
- Calcium signaling with FLIPR Calcium 6 and 6-QF Assay Kits on the FlexStation 3 reader
- HTRF IP-One Gq assay on SpectraMax readers
cAMP Assays (GPCR, Gi/Gs coupled)
Monitoring levels of cAMP, a second messenger produced in response to activation of adenylate cyclase, is one of the most common ways to screen for agonists and antagonists of Gi/Gs protein-coupled receptors. cAMP levels can be monitored using fluorescent molecules that bind to cAMP and are detected using a fluorescence plate reader. Some cAMP assays require other detection modes, such as time-resolved fluorescence.
Here are a few application notes you may find of interest:
cAMP Assays (GPCR, Gi/Gs coupled)
Monitoring levels of cAMP, a second messenger produced in response to activation of adenylate cyclase, is one of the most common ways to screen for agonists and antagonists of Gi/Gs protein-coupled receptors. cAMP levels can be monitored using fluorescent molecules that bind to cAMP and are detected using a fluorescence plate reader. Some cAMP assays require other detection modes, such as time-resolved fluorescence.
Here are a few application notes you may find of interest:
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Kinases, Phosphatases, Phosphodiesterases
For many years, assays of kinase activity were performed using radioactive isotopes or highly specific antibodies. Molecular Devices’ proprietary IMAP® Technology provides a non-radioactive, homogeneous assay amenable to high-throughput screening and applicable to a wide variety of kinases, phosphatases, and phosphodiesterases, regardless of their substrate peptide sequences.
Here are a few application notes you may find of interest:
Kinases, Phosphatases, Phosphodiesterases
For many years, assays of kinase activity were performed using radioactive isotopes or highly specific antibodies. Molecular Devices’ proprietary IMAP® Technology provides a non-radioactive, homogeneous assay amenable to high-throughput screening and applicable to a wide variety of kinases, phosphatases, and phosphodiesterases, regardless of their substrate peptide sequences.
Here are a few application notes you may find of interest:
Neuronal Calcium Flux
Monitoring changes in intracellular calcium is an extremely useful technique for investigating the variety of roles that calcium ions play in functioning neurons. Direct measurement of the dynamic calcium flux within neuronal networks reveals how neurons process signals from the extracellular space.
This application note demonstrates the feasibility of measuring intracellular calcium flux in primary rat cortical neurons in a fluorescent microplate format:
Neuronal Calcium Flux
Monitoring changes in intracellular calcium is an extremely useful technique for investigating the variety of roles that calcium ions play in functioning neurons. Direct measurement of the dynamic calcium flux within neuronal networks reveals how neurons process signals from the extracellular space.
This application note demonstrates the feasibility of measuring intracellular calcium flux in primary rat cortical neurons in a fluorescent microplate format:
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Single and Dual Luciferase Reporter Gene Assays
Reporter gene assays are important tools for studying gene expression associated with the activation of cellular pathways. Cells are transfected with a plasmid containing the reporter gene and a sequence of interest, typically a promoter or other transcriptional control element. When the promoter is activated, the reporter gene is expressed and its levels can be measured.
Learn more about luciferase reporter gene assays with these application notes:
- Monitor NF-kB activation with a sensitive dual reporter assay on the SpectraMax iD5 microplate reader
- Detect dual luciferase expression on the FlexStation 3 microplate reader
- Dual-Luciferase assays in 96- and 384-well formats on the SpectraMax L
- Measuring luciferase expression using the SpectraMax Glo Steady-Luc Reporter Assay Kit
Single and Dual Luciferase Reporter Gene Assays
Reporter gene assays are important tools for studying gene expression associated with the activation of cellular pathways. Cells are transfected with a plasmid containing the reporter gene and a sequence of interest, typically a promoter or other transcriptional control element. When the promoter is activated, the reporter gene is expressed and its levels can be measured.
Learn more about luciferase reporter gene assays with these application notes:
- Monitor NF-kB activation with a sensitive dual reporter assay on the SpectraMax iD5 microplate reader
- Detect dual luciferase expression on the FlexStation 3 microplate reader
- Dual-Luciferase assays in 96- and 384-well formats on the SpectraMax L
- Measuring luciferase expression using the SpectraMax Glo Steady-Luc Reporter Assay Kit
Resources of Cellular Signaling
Application Note
Spectral Fusion™ Illumination technology for an extended dynamic range on the SpectraMax i3x Multi-Mode Microplate Reader
Spectral Fusion™ Illumination technology for an extended dynamic range on the SpectraMax i3x Multi-Mode Microplate Reader
Many microplate readers and spectrophotometers use a photomultiplier tube (PMT) to detect fluorescence and luminescence signals. PMTs work by initially detecting photons from the sample…
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Virology and Vaccine Research Microplate Reader Solutions
Virology and Vaccine Research Microplate Reader Solutions
We understand what a challenging time this is for those who are working to get accelerated testing and medicines to the market as soon as possible and are committed to supporting scientists…
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eBook
The Ultimate Guide to Microplate Reader Solutions
The Ultimate Guide to Microplate Reader Solutions
Evaluating microplate readers doesn’t have to be overwhelming. First, consider your application needs. If your budget is modest, a single-mode reader dedicated to your main application may b…
Publications
Multiple Steps from Microplate-Reader Workstations
Multiple Steps from Microplate-Reader Workstations
Scientists take many roads to overcoming bottlenecks in workflows. For example, a workstation including a microplate reader can handle multiple steps, which improves efficiency and adds spee…
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Application Note
HTRF IP-One Gq assay on SpectraMax Readers
HTRF IP-One Gq assay on SpectraMax Readers
HTRF® is a versatile technology developed by Cisbio Bioassays for detecting biomolecular interactions.
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Application Note
Detect GPCR activity with the cAMP-Gs HiRange HTRF assay
Detect GPCR activity with the cAMP-Gs HiRange HTRF assay
HTRF® is a versatile technology developed by Cisbio for detecting biomolecular interactions.
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Application Note
Calcium flux assay for in vitro neurotoxicity studies and drug screening
Calcium flux assay for in vitro neurotoxicity studies and drug screening
Calcium imaging, or monitoring changes in intracellular calcium, is an extremely useful technique for investigating the variety of roles that calcium ions have in functioning neurons.…
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Application Note
Monitor NF-κB activation with a sensitive dual luciferase reporter assay on the SpectraMax iD5
Monitor NF-κB activation with a sensitive dual luciferase reporter assay on the SpectraMax iD5
Reporter genes are valuable tools for studying gene expression, serving as surrogates for genes involved in various signaling pathways and disease conditions. Luciferases are the most…
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Application Note
Complete cAMP workflow solution using the CatchPoint cAMP Fluorescent Assay Kit
Complete cAMP workflow solution using the CatchPoint cAMP Fluorescent Assay Kit
G-Protein Coupled Receptors (GPCRs) are important transmembrane proteins that translate extracellular signals into intracellular responses. These intracellular responses are comprised of…
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Application Note
HTRF cAMP dynamic 2 and IP-One assays on the SpectraMax M5e reader
HTRF cAMP dynamic 2 and IP-One assays on the SpectraMax M5e reader
HTRF is a versatile technology developed by Cisbio for detecting molecular interactions between biomolecules.1 Combining the features of fluorescence resonance energy transfer (FRET) and…
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Application Note
Detect dual luciferase expression on the FlexStation 3 microplate reader
Detect dual luciferase expression on the FlexStation 3 microplate reader
Reporter gene assays are important tools for studying gene expression associated with the activation of cellular pathways. Cells are transfected with a plasmid containing the reporter gene…
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Application Note
IMAP FP kinase assays on the SpectraMax M5 Multi-Mode Microplate Reader
IMAP FP kinase assays on the SpectraMax M5 Multi-Mode Microplate Reader
Protein kinases are central to the regulation of many cellular processes. In recent years they have emerged as one of the most important classes of drug targets for cancer and many other…
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Application Note
Dual-Luciferase assays in 96- and 384-well formats on the new SpectraMax L
Dual-Luciferase assays in 96- and 384-well formats on the new SpectraMax L
Reporter gene assays are used to study the expression of eukaryotic genes. In dual reporter gene assays cells are transfected with two vectors, the first containing an experimental reporter…
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Application Note
Optimization of a muscarinic M3-receptor assay using frozen CHO cells on the FlexStation 3 reader
Optimization of a muscarinic M3-receptor assay using frozen CHO cells on the FlexStation 3 reader
Cell-based assays can often be challenging and time consuming. To facilitate and streamline this complicated process, frozen cells, which can be assayed without prior cultivation, have…
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Application Note
Calcium signaling with FLIPR Calcium 6 and 6-QF Assay Kits on the FlexStation 3 reader
Calcium signaling with FLIPR Calcium 6 and 6-QF Assay Kits on the FlexStation 3 reader
FLIPR® Calcium Assay Kits from Molecular Devices employ sensitive calcium indicators and proprietary masking dyes to enable researchers to conduct highly sensitive fluorescent screens of G-…
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Application Note
IMAP phosphodiesterase assays on SpectraMax Multi-Mode Microplate Readers
IMAP phosphodiesterase assays on SpectraMax Multi-Mode Microplate Readers
Cyclic nucleotide phosphodiesterases (PDEs) are a group of enzymes that degrade the phosphodiester bond of cAMP and cGMP, second messengers that are involved in a variety of biological…
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Application Note
HTRF cAMP HiRange Assay on SpectraMax Multi-Mode Microplate Readers
HTRF cAMP HiRange Assay on SpectraMax Multi-Mode Microplate Readers
HTRF is a versatile technology developed by Cisbio Bioassays for detecting biomolecular interactions1. It combines fluorescence resonance energy transfer (FRET) technology with time-…
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Application Note
Monitoring Gq protein-coupled receptor activation on the SpectraMax i3x reader with injector module
Monitoring Gq protein-coupled receptor activation on the SpectraMax i3x reader with injector module
Gq protein-coupled receptor activation is commonly monitored in live cells in real time using calcium-sensitive dyes on a fluorescence plate reader. Automated liquid handling within the…
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Application Note
Measuring luciferase expression using the SpectraMax Glo Steady-Luc Reporter Assay Kit
Measuring luciferase expression using the SpectraMax Glo Steady-Luc Reporter Assay Kit
The use of gene reporters such as luciferase permits highly sensitive and nondestructive monitoring of gene expression. Firefly luciferase, a 61 kD monomeric protein, is especially…
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