Phospho-stat Detection
Phospho-stat Detection - Related Videos
10:52
Radiolabeling and Quantification of Cellular Levels of Phosphoinositides by High Performance Liquid Chromatography-coupled Flow Scintillation
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Cited by 21 •
2016
Phosphoinositides are signaling lipids whose relative abundance rapidly changes in response to various stimuli. This article describes a method to measure the abundance of phosphoinositides by metabolically labeling cells with 3H-myo-inositol, followed by extraction and deacylation. Extracted glycero-inositides are then separated by high-performance liquid chromatography and quantified by flow scintillation.
06:23
pH Modulation Assay on Supported Lipid Bilayers to Detect Protein-Phosphoinositide Interactions
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2025
In this video, we demonstrate protein-phosphoinositide interactions in a microfluidic pattern using the pH modulation assay, using pH-sensitive fluorescent dye tagged to phosphatidylethanolamine lipid of a supported lipid bilayer, called a microfluidic platform. The modulations in pH after protein binding to phospholipid cause fluorescence quenching.
11:19
Merging Absolute and Relative Quantitative PCR Data to Quantify STAT3 Splice Variant Transcripts
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Cited by 2 •
2016
Tandem splicing events occur at sites less than 12 nucleotides apart. Quantifying ratios of such splice variants is feasible using an absolute quantitative PCR approach. This manuscript describes how splice variants of the gene STAT3, in which two splicing events results in Serine-701 inclusion/exclusion and α/β C-termini, can be quantified.
01:42
Phosphoinositides and PIPs
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2023
Phosphoinositides are a group of phospholipids containing a glycerol backbone with two fatty acid chains and a phosphate attached to a myoinositol sugar ring. The inositol head group extends into the cytoplasm, where it is modified by adding phosphate groups to form phosphatidylinositol phosphates or PIPs. Different phosphoinositides are synthesized and recruited on the cytosolic face of the plasma membrane. The localization of specific phosphoinositides concentrated in separate membrane...
10:58
PIP-on-a-chip: A Label-free Study of Protein-phosphoinositide Interactions
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Cited by 7 •
2017
Here we present a supported lipid bilayer in the context of a microfluidic platform to study protein-phosphoinositide interactions using a label-free method based on pH modulation.
FAQs
Phosphorylation connects receptor-associated kinase activity with STAT-dependent gene regulation. After cytokine, growth factor, or other stimulation, JAK-mediated modification supports STAT dimerization and nuclear entry, allowing the proteins to influence transcription. Measuring the phosphorylated form therefore links an upstream biochemical event to the signaling state that precedes changes in gene expression.
The phosphorylated STAT species provides a direct biochemical readout of signaling activity rather than relying only on later gene-expression changes. Its measurement can reveal whether a stimulus has engaged the pathway and whether that response differs between treatments. This makes phosphorylation status useful for examining signaling changes in both experimental and disease-related contexts.
Interpretation depends on the relationship among the stimulus, receptor-associated kinases, STAT proteins, and their phosphorylation state. Cytokines and growth factors can initiate signaling, while JAK activity contributes to STAT modification. The detected phospho-STAT signal should therefore be considered in the context of the treatment or stimulus used and the specific STAT-related pathway being investigated.
Phospho-specific antibodies support several measurement formats, including Western blotting, flow cytometry, and immunofluorescence. Western blotting can assess phosphorylated proteins in biochemical samples, whereas flow cytometry and immunofluorescence enable detection in cell-based formats. The appropriate choice depends on whether the experiment emphasizes biochemical measurement, cellular analysis, or visualization of signaling-related staining.
A typical workflow links a defined cellular stimulus or treatment to antibody-based measurement of the resulting phosphorylated STAT signal. Researchers select a detection format, apply phospho-specific antibodies, and compare the observed signal across treatments or conditions. This design allows pathway activation or suppression to be evaluated in relation to the experimental stimulus.
This approach is used to characterize pathway activation, compare cellular responses to different treatments, and investigate signaling changes associated with disease. It also supports evaluation of inhibitors directed at JAK-STAT and related biochemical pathways. By measuring a signaling intermediate, researchers can connect treatment effects with changes occurring within the pathway rather than considering only downstream outcomes.