8 Cell Lysis Methods to Break Cell Walls
Choosing the right cell lysis method is critical for success. Some methods, such as high-temperature lysis and sonication, may denature samples. French presses and rotor-stators are more gentle but expensive, while freeze–thawing is gentle but slow. Enzymatic and chemical lysis methods are gentle, but the added species may need to be removed or destroyed before downstream experiments.
We all need to lyse cells to extract the goodness—our samples—from them.
But there are many cell lysis methods. Some are harsh, while some are gentle. Some are laborious, while some are easy. Some require dedicated equipment, while some do not. So which one do you choose?
The best choice is probably the one that requires the least work that doesn’t harm your sample.
With that in mind, here’s a brief insight into eight cell lysis methods for your experiments.
8 Cell Lysis Methods Explained
The cell lysis methods below fall into two classes:
- Mechanical methods.
- Non-mechanical methods.
Since non-mechanical methods involve adding chemicals or enzymes to the cells, it’s fairly clear which class each lysis method belongs to.
The advantage of mechanical methods is that they are direct, and you have more control over them. But some mechanical methods are harsh and may denature your sample if not performed diligently.
The advantage of non-mechanical methods is that chemicals or enzymes lysis the cells for you. But you may have to remove these later to prevent them from interfering with your experiments.
Let’s get into it.
1. Mortar and Pestle
This method involves giving the cells a good old grinding. Note that the cells do not have to be suspended in anything for this method to work.
This cell lysis method is often used for plant samples frozen in liquid nitrogen since the specimens are comparatively bulky.
Also, when you’ve lysed the cells, metabolites and samples may be extracted by adding solvents.
And you don’t need any specialist kit—just the mortar and pestle (Figure 1)!
2. Bead Beating
Glass or ceramic beads can crack open cells. It might not sound like it, but this kind of mechanical shear is gentle enough to keep organelles intact.
It can be used with all kinds of cells, just add beads to a suitable amount of cell suspension and vortex!
Alternatively, slightly more exotic equipment, such as bead mills, can standardize your cell lysis. So think about what will work for you.
This is perhaps the most common cell lysis method, and most readers will be familiar with it.
Ultrasonic homogenizers (usually just called sonicators) work by inducing rapid vibrations in a titanium probe immersed in the cell solution.
This causes cavitation, a process in which tiny bubbles are produced within the cell suspension. When these bubbles explode, a local shockwave disrupts the cell wall, just like an explosion disrupts things in the blast radius. Only the medium is liquid, not air.
This method is very popular for plant and fungal cells and is the method of choice for protein purification.
While sonication is automated—you just set the timer and go—it has several disadvantages.
It generates a lot of heat, and you must be careful to avoid heating your sample. For this reason, sonication is usually done on ice and in pulses.
This method is also extremely loud. So the process is usually done in a soundproof cabinet or with ear defenders.
Sonication is also notorious for creating aerosols (tiny droplets that waft through the air and that you can breathe in). So don’t sonicate any infectious material.
And you will need a sonicator. The good news is they are really common and have interchangeable tips to suit different sample sizes.
Homogenizers use shearing forces to lyse cells, similar to the bead-beating method.
Homogenization can be performed by squeezing cells through a tube that is slightly smaller than the cell dimensions, thereby shearing away the outer layer (how a French press lyses cells) or by using a rotating blade to lyse cells, just like a kitchen blender (how a rotor-stator lyses cells).
Both are comparatively gentle, and people who purify membrane proteins prefer the French press cell lysis method. Plus, French presses can break the rock-hard plasma membrane.
But French presses are relatively cumbersome, require greasing and maintenance, and can clog easily.
Rotor-stators, on the other hand, are quick and efficient, but you can usually only process a few tens of microliters of cells at once.
Plus, French presses and rotor-stators may cost a few thousand dollars.
If you lyse cells using these bits of kit, beware of cross-contamination and note that you can add enzymes such as lysozyme to help the process.
5. Freeze–Thaw Lysis
This one’s as simple as it sounds.
The freeze–thaw cell lysis method creates ice crystals (remember, water expands when it freezes) that melt when the sample is thawed. Several freeze–thaw cycles ultimately rupture the cells.
It’s comparatively gentle and, by definition, doesn’t generate any heat. But it’s time-consuming.
You can speed things up by using liquid nitrogen, but even so, it will take a while.
6. High-Temperature Lysis
High temperatures and pressures break the chemical bonds within cell walls—but also denature proteins.
So it’s a quick and dirty cell lysis method. You can use a microwave or autoclave to generate the heat (and pressure).
But it goes without saying that you should avoid this method if your sample is denatured by heat.
7. Enzymatic Lysis
There are enzymes that can destroy the cell wall for you!
The enzymatic cell lysis method breaks open cells by using enzymes that degrade cell walls and membranes.
Simply incubate a suspension of your cells with an appropriate enzyme or combination of enzymes, which digest the cell wall components.
Depending on what organism you work with, you can use cellulases, chitinase, bacteriolytic enzymes like lysozyme (it destroys peptidoglycans), mannanase, or glycanase, etc.
This method is also great if you want to isolate the protoplast (a cell without the wall).
And it’s really quite gentle, meaning you don’t have to worry about denaturing your sample. But you might have to remove or destroy the enzymes later on.
8. Chemical Lysis
Organic solvents like alcohols, ether, or chloroform can disrupt the cell wall by permeabilizing cell walls and membranes.
This method is especially handy if you want to extract hydrophobic molecules like plant pigments because you can pick a solvent that will extract them.
And you can combine this method with one that uses shearing forces to maximize the fraction of cells lysed.
And note also, you can use EDTA specifically to disrupt the cell walls of Gram-negative bacteria, whose cell walls contain lipopolysaccharides that are stabilized by cations like Mg2+ and Ca2+.
EDTA will chelate the cations leaving holes in the cell walls, and you can dialyze the EDTA away later!
Extracting What You Need: Cell Lysis Methods Summarized
So which cell lysis method works best for you?
Now you are armed with the knowledge to burst your cells in a way that works for you. From methods that seem more like preparing dinner to fancy French presses, and everything in between.
For extra information to help you pick a lysis method, check out our article on the different types of cell walls.
And if we’ve missed your favorite cell lysis method or some other crucial fact, let us know in the comments section.
Originally published July 2013. Revised and updated April 2023.
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I want to break the cell wall in a suspension containing yeast and bacteria to extract the internal metabolites especially vitamins. Which of the mentioned methods is more useful? the capacity of the suspension is 500 lit in each bach.
Why we need to break down the cell mechanically? Thanks.
I wanted to know the chemical which can be used for clearing the cytoplasm for screening the chromosomes for karyotype studies in connection with orchids
The purpose of cell disruption is to obtain internal substances, like chromosome. do you have a way to get chromosome without breakage