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Cell lysis 101: 8 methods to break down cell walls

In Part 1, I introduced the types of cell walls out there and what they are made of – now it’s time to learn how to get through them.

1.  Mechanical methods of cell disruption

Mechanical cell disruption is really just that: forcing open the cell wall and spilling the contents. The advantage to mechanical disruption is that no chemicals are introduced that might interfere with the substance you want to extract. The drawback however is that the method has to be carefully adjusted as to not destroy that molecule of interest.

Mortar and Pestle

Just give the cells a good old grinding. This does not have to be in suspension and is often done with plant samples frozen in liquid nitrogen. When the material has been disrupted, metabolites can be extracted by adding solvents.

Beadbeating

Glass or ceramic beads are used to 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 an equal amount of cell suspension and vortex!

Ultrasonictor “Amplichron®” by LKB instruments.

Sonication

Ultrasonic homogenizers work by inducing vibration in a titanium probe that is immersed in the cell solution. A process called caviation occurs, in which tiny bubbles are formed and explode, producing a local shockwave and disrupting cell walls by pressure change. This method is very popular for plant and fungal cells but comes at a disadvantage: It’s very loud and has to be performed in an extra room, otherwise you will be very unpopular.

Homogenizer

Homogenizers use shearing forces on the cell similar to the bead method. Homogenization can be performed by squeezing cells through a tube that is slightly smaller than them, thereby shearing away the outer layer (French Press) or by using a rotating blade like in a blender (Rotor-Stator Processors).

Freezing

Freeze-thaw cycles work by formation of ice crystals and cell expansion upon thawing, ultimately leading to rupture. Used for algae and soft plant material. The drawback is that it is very time-consuming.

High temperatures (Microwave, Autoclave)

High temperatures (and pressure) disrupt bonds within cell walls, but also denature proteins. Although it is quick, you better find another method if your application is affected by the damage heat does to the rest of the cell.

2.  Non-mechanical methods

Non-mechanical methods involve the addition of enzymes or chemicals that specifically break down cell wall components.  They are often used in combination with mechanical force to ensure complete disruption of the cell. The disadvantage to their use is that they often have to be removed from the sample afterwards.

Enzymes

Naturally occurring enzymes can be used to remove the cell wall specifically, for example when isolating the protoplast (cell without the wall). Depending on what organism you work with, that can be cellulases, chitinase, bacteriolytic enzymes like lysozyme (destroys peptidoglycans), mannase, glycanase (etc. ).

Chemicals

Organic solvents like alcohols, ether or chloroform can disrupt the cell wall by permeabilizing cell walls and membranes. They are especially handy if you want to extract hydrophobic molecules (like plant pigments) because they will be collected in the solvent. Often used on plants in combination with shearing forces.

EDTA can be used 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.

Now you are armed with ideas and ready to attack your cells! If you want to go back to finding out what your cell wall is likely to consist of before you chose a method, read it here!

8 Comments

  1. Neelima on June 15, 2018 at 3:24 pm

    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

  2. Yuanchun King on December 22, 2017 at 2:09 pm

    The purpose of cell disruption is to obtain internal substances, like chromosome. do you have a way to get chromosome without breakage

  3. Chloie A on July 16, 2017 at 12:51 pm

    This is helpful, which method would be most suitable for a spinach leaf (with the leaf still intact after the cell wall is broken)?

  4. Jason Hirsch on February 11, 2017 at 11:23 pm

    I’d like to thank you for these ideas- quick, concise, with all associated positives and negatives. This has given me a boost to start reading other ideas.

  5. Gina on January 9, 2016 at 3:01 am

    If a fungal enzyme is used to break cell walls in plants, would that extracted fungal enzyme break cell walls in humans if placed in a supplement?

    • Laura-Nadine Schuhmacher on May 30, 2018 at 9:57 am

      human cells don’t have cell walls 🙂 if you were using an enzyme like that in a supplement, it would be to help humans break down plant cell walls, for example if we were eating grass.

  6. Iherb on November 29, 2015 at 8:02 pm

    Thank you for this interesting post! What would be the recommended method for tree pollen?

  7. iva gyurgina on July 16, 2015 at 5:13 pm

    Thank you for the interesting blog post on different methods of cell disruption. You mentioned that one disadvantage for the sonication method is the level of noise. One thing to keep in mind is that you can install a sound abating enclosure (some prefer to just use noise canceling headphones). Also, if you are interested in that cell disruption method you can check out our blog where we regularly post ultrasonic equipment particles:
    http://sonomechanics.com/applications/pharmaceutical/cell_disruption/
    http://blog.sonomechanics.com/blog

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