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Permeabilizing cells: some methods and applications for the study of intracellular processes
Methods in Enzymology
|January 1, 1990
Summary
Controlled cell permeabilization techniques, including electropermeabilization and detergent methods, allow researchers to access and manipulate intracellular events. These methods are crucial for studying stimulus-response coupling in various cell types.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Controlled permeabilization of plasma membranes is essential for accessing the cell interior and manipulating intracellular events.
- Understanding stimulus-response coupling requires precise control over cellular compartments.
Purpose of the Study:
- To review and compare common cell permeabilization techniques.
- To highlight the utility of permeabilized cells in studying cellular mechanisms like exocytosis.
Main Methods:
- Electropermeabilization creates small pores (~2 nm).
- Permeabilization with detergents (saponin, digitonin) or pore-forming toxins (alpha-toxin, streptolysin O) creates larger pores.
- Pore size determines the ability to introduce molecules of varying sizes.
Main Results:
- Different permeabilization methods yield varying pore sizes, impacting molecular entry.
- Loss of cytosolic constituents can be a disadvantage of larger pore methods.
- Exocytosis in secretory cells is often triggered by Ca2+, but requirements vary across cell types.
Conclusions:
- Permeabilized cells serve as an intermediate model between intact cells and isolated organelles.
- These preparations offer significant opportunities to advance the understanding of stimulus-response coupling mechanisms.
- Cell-specific nucleotide, anion, and cation requirements are critical for exocytosis.