Digitonin
Digitonin - Related Videos
14:39
Cholinergic Ligand–dependent Modulation of Oxidative Phosphorylation Coupling in Digitonin-permeabilized BE(2)-C Neuroblastoma Cells
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2026
We present a protocol to selectively permeabilize plasma membrane, enabling targeted delivery of substrates and inhibitors to assess mitochondrial oxidative phosphorylation (OXPHOS) in cultured cells, overcoming plasma membrane transport barriers that typically hinder such analysis. Using this technique, we demonstrated how OXPHOS function is modulated by muscarinic agonists and antagonists.
06:27
An Assay to Quantify Cytosolic and Vacuolar Salmonella in Infected Primary Macrophages
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2025
This video demonstrates an assay to quantify cytosolic and vacuolar Salmonella typhimurium inside bone marrow-derived macrophages using differential permeabilization. The macrophages are infected with a fluorescent protein-expressing Salmonella, which is internalized inside a specialized vacuolar structure called Salmonella-containing vacuole or SCV. A subset of bacteria escape the SCV and enter the cytosol. The cholesterol-rich plasma membrane of macrophages is permeabilized using digitonin,...
08:43
Simultaneous Measurement of Mitochondrial Calcium and Mitochondrial Membrane Potential in Live Cells by Fluorescent Microscopy
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Cited by 29 •
2017
Mitochondria can utilize the electrochemical potential across their inner membrane (ΔΨm) to sequester calcium (Ca2+), allowing them to shape cytosolic Ca2+ signaling within the cell. We describe a method for simultaneously measuring mitochondria Ca2+ uptake and ΔΨm in live cells using fluorescent dyes and confocal microscopy.
05:47
Oxygen Flux Measurements to Evaluate Mitochondrial Respiration in Living and Permeabilized Cells
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2026
Source: Elettra Leo1, Luiza H.D. Cardoso1, Sabine Schmitt1, Erich Gnaiger11Oroboros InstrumentsThis video explains how oxygen flux is used to assess mitochondrial respiration in living and permeabilized mammalian cells. It demonstrates the sequential use of digitonin, metabolic substrates, ADP, an uncoupler, and an inhibitor to characterize different respiratory states. The resulting oxygen flux profile reveals mitochondrial function and energy-coupling efficiency under defined conditions.
11:32
Mapping RNA-RNA Interactions Globally Using Biotinylated Psoralen
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Cited by 20 •
2017
Here, we detail the method of Sequencing of Psoralen crosslinked, Ligated, and Selected Hybrids (SPLASH), which enables genome-wide mapping of intramolecular and intermolecular RNA-RNA interactions in vivo. SPLASH can be applied to study RNA interactomes of organisms including yeast, bacteria and humans.
FAQs
Digitonin preferentially affects membranes containing cholesterol because its membrane-disrupting action depends on binding that sterol and forming complexes. The plasma membrane is therefore made accessible while many cholesterol-poor organelle membranes can remain comparatively intact. This distinction lets investigators expose intracellular contents without treating every cellular membrane as equally vulnerable, preserving compartmental information for subsequent analysis.
Concentration and exposure time determine how extensively digitonin disrupts the plasma membrane. Carefully controlled values can provide intracellular access while preserving many organelle membranes, whereas less controlled treatment may compromise cellular architecture. Adjusting these two variables allows researchers to tailor permeabilization to the intended analysis, such as cytosolic protein release or examination of compartment-specific processes.
Selective disruption preserves differences between the plasma membrane and intracellular organelle membranes. That preservation matters because researchers can access cellular contents while retaining information about where components originated and which compartments remain intact. As a result, digitonin treatment supports investigations of membrane organization and compartment-specific biology rather than reducing the cell to an undifferentiated mixture.
Researchers should deliberately control both the digitonin concentration and the duration of exposure, then assess whether the treatment provides the needed intracellular access without completely destroying cellular architecture. This balance is central to the method: insufficient treatment may limit access, while excessive disruption can reduce compartmental information. The chosen conditions should therefore match the planned biological analysis.
Digitonin-permeabilized cells can support immunofluorescence, cell fractionation, and assays of mitochondrial function or cytosolic protein release. In immunofluorescence, intracellular components become accessible for analysis. In fractionation and release assays, selective membrane disruption helps examine cellular components or soluble contents in relation to their compartments, extending the technique beyond simple membrane perturbation.
For cell fractionation, controlled permeabilization can help isolate or examine cellular components while retaining distinctions between membrane compartments. In mitochondrial studies, preserving many cholesterol-poor organelle membranes allows mitochondrial function to be assayed after the plasma membrane has been made accessible. These uses connect digitonin’s membrane selectivity to practical analysis of intracellular organization and activity.