Intracellular Protein Detection
Intracellular Protein Detection - Related Videos
04:58
Isolation of Cell-Surface and Intracellular Proteins from an Astrocyte Culture
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2025
This video demonstrates the isolation of cell-surface and intracellular proteins from an astrocyte culture. The astrocyte culture is placed on ice to inhibit endocytosis, and then the cell-surface proteins are labeled with a biotinylation reagent. The cells are lysed to release the biotinylated cell-surface proteins and non-biotinylated intracellular proteins. Finally, streptavidin-coated beads are used to separate the biotinylated cell-surface proteins from the intracellular proteins.
10:08
Purification and Aggregation of the Amyloid Precursor Protein Intracellular Domain
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Cited by 1 •
2012
A method for large-scale purification of the APP intracellular domain (AICD) is described. We also describe methodology to induce in vitro AICD aggregation and visualization by atomic force microscopy. The methods described are useful for biochemical/structural characterization of the AICD and the effects of molecular chaperones on its aggregation.
06:15
Biotinylated Cell-Penetrating Peptide Probe to Detect Protein-Protein Interactions
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2025
This video demonstrates a technique that uses biotinylated cell-penetrating peptides to detect intracellular protein-protein interactions. The cell-penetrating peptides facilitate the conjugated peptide to enter a cell and specifically bind to its interacting proteins. The biotin, after binding to avidin, allows purification of the interacting proteins’ complex from a cell lysate.
09:00
Detection of Protein Ubiquitination
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Cited by 96 •
2009
Ubiquitination is a key posttranslational modification carried out by a set of three enzymes. Mutations of genes involved in this modification are associated with many different human diseases. Here, we describe protocols to detect protein ubiquitination in cultured cells in vivo and test tubes in vitro.
03:40
Evaluation of Stromal Cytokine-Induced Intracellular Protein Phosphorylation Using Phospho-Flow Cytometry
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2025
This video demonstrates the evaluation of stromal cytokine-induced intracellular phosphorylation of proteins in leukemia cells. These intracellular phosphorylated proteins are quantified using the phospho-flow cytometry technique.
FAQs
The plasma membrane restricts access to proteins located inside cells, so fixation and permeabilization prepare cells for detection. Fixation preserves cellular structure, while permeabilization allows labeled antibodies or other detection reagents to reach intracellular targets. These steps help retain information about protein location and distribution, including differences between subcellular compartments.
Labeled antibodies detect a protein after cells have been prepared for access, whereas genetically encoded fluorescent tags mark the protein through an attached fluorescent signal produced within the cell. Both approaches support intracellular localization and measurement, but they provide different routes to generating detectable signal for microscopy or flow cytometry.
The readout depends on the information needed from the experiment. Microscopy records where signal appears, making it useful for examining intracellular distribution and subcellular compartments. Flow cytometry measures signal in individual cells, supporting comparisons of protein presence or amount across many cells. Both approaches preserve cell-level information while emphasizing different types of outcomes.
A typical workflow prepares cells by fixation and permeabilization, exposes intracellular targets to labeled antibodies or genetically encoded fluorescent tags, and then records the resulting signal. The selected readout, microscopy or flow cytometry, determines how the measurement is captured. This sequence connects cellular preparation with analysis of protein amount, location, or distribution.
These measurements can reveal how proteins move through cells, where they accumulate, and how their expression changes. They can also help characterize signaling pathways, disease mechanisms, and cellular identity. Because detection retains information from individual cells and compartments, researchers can relate protein patterns to specific cellular states rather than relying only on a population-wide summary.
Researchers can compare intracellular protein signals after cells encounter drugs or environmental conditions. Changes in protein amount, distribution, or localization provide evidence that the cellular response has altered expression or trafficking. Microscopy can show where those changes occur, while flow cytometry can compare responses among individual cells, supporting studies of biological variability.