Cell disruption by nitrogen decompression
Encyclopedia
Cell disruption by rapid decompression is one of several methods of cell disruption
and is also called explosive decompression
or cell bomb.
According to manufacturers of nitrogen decompression devices, the method is particularly well suited for treating mammalian and other membrane bound cells. It has also been used successfully for treating plant cell
s, for releasing virus from fertilized eggs and for treating fragile bacteria. It is not recommended for untreated bacterial cells. Yeast
, fungus
, spores and other materials with tough cell walls do not respond well to this method.
. Then, when the gas pressure is suddenly released, the nitrogen comes out of the solution as expanding bubbles that stretch the membranes of each cell until they rupture and release the contents of the cell.
Nitrogen decompression is claimed to be more protective of enzyme
s and organelle
s than ultrasonic
and mechanical homogenizing methods and to compare favorably to the controlled disruptive action obtained in a PTFE and glass mortar and pestle homogenizer. While other disruptive methods depend upon friction or a mechanical shear
ing action that generate heat, the nitrogen decompression procedure is accompanied by an adiabatic expansion that cools the sample instead of heating it.
The blanket of inert nitrogen gas that saturates the cell suspension and the homogenate offers protection against oxidation of cell components. Although other gases: carbon dioxide
, nitrous oxide
, carbon monoxide
and compressed air have been used in this technique, nitrogen
is preferred because of its non-reactive nature and because it does not alter the pH of the suspending medium. In addition, nitrogen is preferred because it is generally available at low cost and at pressures suitable for this procedure.
Once released, subcellular substances are not exposed to continued attrition
that might denature
the sample or produce unwanted damage. There is no need to watch for a peak between enzyme activity and percent disruption. Since nitrogen bubbles are generated within each cell, the same disruptive force is applied uniformly throughout the sample, thus ensuring unusual uniformity in the product. Cell-free homogenates can be produced.
Cell disruption
Cell disruption is a method or process for releasing biological molecules from inside a cell.- Choice of disruption method:The production of biologically-interesting molecules using cloning and culturing methods allows the study and manufacture of relevant molecules.Except for excreted molecules,...
and is also called explosive decompression
Explosive decompression
Uncontrolled decompression refers to an unplanned drop in the pressure of a sealed system, such as an aircraft cabin and typically results from human error, material fatigue, engineering failure or impact causing a pressure vessel to vent into its lower-pressure surroundings or fail to pressurize...
or cell bomb.
Applications
The technique is used to:- HomogenizeHomogenization (biology)Homogenization is a process that involves breaking apart cells — releasing organelles and cytoplasm.When the purpose is to extract organelles, it is frequently done in two steps; first using a blender to break the tissue up, and then with an ultrasonic or mechanical tissue disruptor. The...
cells and tissues - Release intact organelles
- Prepare cell membraneCell membraneThe cell membrane or plasma membrane is a biological membrane that separates the interior of all cells from the outside environment. The cell membrane is selectively permeable to ions and organic molecules and controls the movement of substances in and out of cells. It basically protects the cell...
s - Release labile biochemicals
- Produce uniform and repeatable homogenates without subjecting the sample to extreme chemical or physical stress.
According to manufacturers of nitrogen decompression devices, the method is particularly well suited for treating mammalian and other membrane bound cells. It has also been used successfully for treating plant cell
Plant cell
Plant cells are eukaryotic cells that differ in several key respects from the cells of other eukaryotic organisms. Their distinctive features include:...
s, for releasing virus from fertilized eggs and for treating fragile bacteria. It is not recommended for untreated bacterial cells. Yeast
Yeast
Yeasts are eukaryotic micro-organisms classified in the kingdom Fungi, with 1,500 species currently described estimated to be only 1% of all fungal species. Most reproduce asexually by mitosis, and many do so by an asymmetric division process called budding...
, fungus
Fungus
A fungus is a member of a large group of eukaryotic organisms that includes microorganisms such as yeasts and molds , as well as the more familiar mushrooms. These organisms are classified as a kingdom, Fungi, which is separate from plants, animals, and bacteria...
, spores and other materials with tough cell walls do not respond well to this method.
How it works
Large quantities of nitrogen are first dissolved in the cell under high pressure within a suitable pressure vesselPressure vessel
A pressure vessel is a closed container designed to hold gases or liquids at a pressure substantially different from the ambient pressure.The pressure differential is dangerous and many fatal accidents have occurred in the history of their development and operation. Consequently, their design,...
. Then, when the gas pressure is suddenly released, the nitrogen comes out of the solution as expanding bubbles that stretch the membranes of each cell until they rupture and release the contents of the cell.
Nitrogen decompression is claimed to be more protective of enzyme
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...
s and organelle
Organelle
In cell biology, an organelle is a specialized subunit within a cell that has a specific function, and is usually separately enclosed within its own lipid bilayer....
s than ultrasonic
Ultrasound
Ultrasound is cyclic sound pressure with a frequency greater than the upper limit of human hearing. Ultrasound is thus not separated from "normal" sound based on differences in physical properties, only the fact that humans cannot hear it. Although this limit varies from person to person, it is...
and mechanical homogenizing methods and to compare favorably to the controlled disruptive action obtained in a PTFE and glass mortar and pestle homogenizer. While other disruptive methods depend upon friction or a mechanical shear
Shearing (physics)
Shearing in continuum mechanics refers to the occurrence of a shear strain, which is a deformation of a material substance in which parallel internal surfaces slide past one another. It is induced by a shear stress in the material...
ing action that generate heat, the nitrogen decompression procedure is accompanied by an adiabatic expansion that cools the sample instead of heating it.
The blanket of inert nitrogen gas that saturates the cell suspension and the homogenate offers protection against oxidation of cell components. Although other gases: carbon dioxide
Carbon dioxide
Carbon dioxide is a naturally occurring chemical compound composed of two oxygen atoms covalently bonded to a single carbon atom...
, nitrous oxide
Nitrous oxide
Nitrous oxide, commonly known as laughing gas or sweet air, is a chemical compound with the formula . It is an oxide of nitrogen. At room temperature, it is a colorless non-flammable gas, with a slightly sweet odor and taste. It is used in surgery and dentistry for its anesthetic and analgesic...
, carbon monoxide
Carbon monoxide
Carbon monoxide , also called carbonous oxide, is a colorless, odorless, and tasteless gas that is slightly lighter than air. It is highly toxic to humans and animals in higher quantities, although it is also produced in normal animal metabolism in low quantities, and is thought to have some normal...
and compressed air have been used in this technique, nitrogen
Nitrogen
Nitrogen is a chemical element that has the symbol N, atomic number of 7 and atomic mass 14.00674 u. Elemental nitrogen is a colorless, odorless, tasteless, and mostly inert diatomic gas at standard conditions, constituting 78.08% by volume of Earth's atmosphere...
is preferred because of its non-reactive nature and because it does not alter the pH of the suspending medium. In addition, nitrogen is preferred because it is generally available at low cost and at pressures suitable for this procedure.
Once released, subcellular substances are not exposed to continued attrition
Attrition
Attrition may refer to:*Physical wear*Attrition warfare, the military strategy of wearing down the enemy by continual losses in personnel and material*Loss of personnel by Withdrawal...
that might denature
Denaturation (biochemistry)
Denaturation is a process in which proteins or nucleic acids lose their tertiary structure and secondary structure by application of some external stress or compound, such as a strong acid or base, a concentrated inorganic salt, an organic solvent , or heat...
the sample or produce unwanted damage. There is no need to watch for a peak between enzyme activity and percent disruption. Since nitrogen bubbles are generated within each cell, the same disruptive force is applied uniformly throughout the sample, thus ensuring unusual uniformity in the product. Cell-free homogenates can be produced.