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5 Laboratory Sterilisation Methods

Effective sterilisation techniques are essential for working with isolated cell lines. For obvious reasons, you don’t want bugs from the environment growing in your nice culture medium, and equally, cultures must be sterilised before disposal. Unless you are using microbiology microscopes to view your cultures, you may be clueless about any unwanted microbe guests. Time to buff up on the various methods of sterilisation, and how do they work to keep your cells healthy, happy and bug-free.

WET HEAT (Autoclaving)

The method of choice for sterilisation in most labs is autoclaving; using pressurised steam to heat the material to be sterilised. This is a very effective method that kills all microbes, spores and viruses, although, for some specific bugs, especially high temperatures or incubation times are required.

Autoclaving kills microbes by hydrolysis and coagulation of cellular proteins, which is efficiently achieved by intense heat in the presence of water.

The intense heat comes from the steam. Pressurised steam has a high latent heat; at 100oC it holds 7 times more heat than water at the same temperature. This heat is liberated upon contact with the cooler surface of the material to be sterilised, allowing rapid delivery of heat and good penetration of dense materials.

At these temperatures, water does a great job of hydrolysing proteins… so those bugs don’t stand a chance.

DRY HEAT (Flaming, baking)

Dry heating has one crucial difference from autoclaving. You’ve guessed it – there’s no water, so protein hydrolysis can’t take place.

Instead, dry heat tends to kill microbes by oxidation of cellular components. This requires more energy than protein hydrolysis so higher temperatures are required for efficient sterilization by dry heat.

For example, sterilisation can normally be achieved in 15 minutes by autoclaving at 121degC, whereas dry heating would generally need a temperature of 160degC to sterilize in a similar amount of time.

FILTRATION

Filtration is a great way to quickly sterilize solutions without heating. Filters, of course, work by passing the solution through a filter with a pore diameter that is too small for microbes to pass through.

Filters can be scintered glass funnels made from heat-fused glass particles or, more commonly these days, membrane filters made from cellulose esters. For removal of bacteria, filters with an average pore diameter of 0.2um is normally used.

But remember, viruses and phage can pass through these filters so filtration is not a good option if these are a concern.

SOLVENTS

Ethanol is commonly used as a disinfectant, although since isopropanol is a better solvent for fat it is probably a better option.

Both work by denaturing proteins through a process that requires water, so they must be diluted to 60-90% in water to be effective.

Again, it’s important to remember that although ethanol and IPA are good at killing microbial cells, they have no effect on spores.

RADIATION

UV, x-rays and gamma rays are all types of electromagnetic radiation that have profoundly damaging effects on DNA, so make excellent tools for sterilization.

The main difference between them, in terms of their effectiveness, is their penetration.

UV has limited penetration in air so sterilisation only occurs in a fairly small area around the lamp. However, it is relatively safe and is quite useful for sterilising small areas, like laminar flow hoods.

X-rays and gamma rays are far more penetrating, which makes them more dangerous but very effective for large scale cold sterilization of plastic items (e.g. syringes) during manufacturing.

So those are some of the main methods for sterilization I can think of. If I’ve missed any, please feel free to let me know in the comments section.

Image Credit: CdePaz

49 Comments

  1. Alan Oluocy on March 28, 2020 at 8:52 pm

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  2. Frank on September 24, 2019 at 1:24 pm

    This is a nice article on Laboratory Sterilisation Methods. It should be shared broadly. Thank you for sharing

  3. syed Tabrez Irfan on July 18, 2019 at 8:46 am

    Please share with me the harmful eggects of Formaldehyde gas if used in Tissue culyure lab for disinfection , specially its effect on Tissue culture plants inside tube. This will be very useful for my work

  4. Surya on May 16, 2019 at 4:01 pm

    My exam? Is what is sterilization. I will use this answer

  5. Sajid on May 13, 2019 at 5:24 am

    I am working to minimize the use of energy as autoclave took longer and use energy a lot In heating and cooling process. Can I use water from ro plant with sensitive membrane to be used for bacterial culturing in lab. Plz guide

  6. Sathya on November 18, 2018 at 8:48 am

    can i use some other chemicals to sterilization

    • Shawn on April 8, 2019 at 12:59 pm

      Yes! Chemicals Used for sterilisation are
      1. Phenol Derivatives : Phenol, Cresol, resorcinol, chloroxylenol
      2. Oxidizing agents :Pot.Permanganate, Hydrogen Peroxide,Benzoyol
      Peroxide
      3. Halogens : Iodine, chlorine
      4. Biguanide : Chlorhexidine
      5. Quarternary Ammonium (Cationic) : Cetrimide, Zephiran
      6. Alcohols : Ethanol, Isopropanol.
      7. Aldehydes : Formaldehyde, Glutaraldehyde
      8. Acids : Boric acid, acetic acid
      9. Metallic salts ; Silver Nitrate, Zince Sulfate, Zinc Oxide, calamine,
      10. Dyes : Gentian violet, proflamine, Acriflamine
      11. Furan derivatives : Nitro flurazone

  7. Rohit on October 21, 2018 at 6:12 am

    What is use in filtration for sterilization

    • Raj Tyagi on September 6, 2019 at 8:23 am

      Heat sensitive samples are sterile with the help of filtration for ex membranes are used in filter sterilization..
      *Liquid or gas samples are sterile by filtration

  8. Dare on September 21, 2018 at 10:53 am

    This is very nice.
    The simple and straight forward classification made everything complete!!
    However, the indicator s of achieving sterilization should have been added.
    Great job!!

  9. nisha makwana on August 22, 2018 at 10:47 am

    its very help full

    • preeti patil on September 5, 2018 at 9:58 am

      Relly

    • ADAM KWAJA on September 8, 2018 at 8:52 am

      it is life bacteria

    • Korirkiprotich on April 26, 2019 at 2:48 pm

      Yu have missed gas sterilization

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