How to Spore-Test an Autoclave

Written by samuel sohlden
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How to Spore-Test an Autoclave
Testing autoclaves ensures that they provide adequate sterilisation. (Ready for surgery image by DGGallery from

Autoclaving is a routine, effective method for sterilising medical instruments, biological media and vaccines. However, autoclaves can malfunction or perform suboptimally for a multitude of reasons, resulting in incomplete sterilisation. Therefore, it's necessary to test autoclaves regularly with Geobacillus stearothermophilus, which is one of the most heat-tolerant species of bacteria. If sterilisation in an autoclave does not destroy the Geobacillus spores, the device is not working properly.

Skill level:

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Things you need

  • Autoclave
  • Spore test
  • Incubator
  • Agar plates

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  1. 1

    Read the directions on the specific spore test product you're using. Some products require you to take certain steps before autoclaving.

  2. 2

    Place the spore test into the autoclave and sterilise for one hour, at standard sterilisation temperature and pressure.

  3. 3

    Wait for the autoclave to depressurise and cool down, and then remove the spore test.

  4. 4

    Spread the autoclaved spore test onto an agar plate, and place it into an incubator for 3 to 48 hours (or follow the manufacturer's instructions). If you don't have an incubator, buy a spore test that you can ship to the manufacturer for incubation.

  5. 5

    Check the agar plates for bacterial growth within the incubation time frame specified by the spore test manufacturer, or wait for results from the manufacturing company.

Tips and warnings

  • You must sterilise the spore test under normal autoclave conditions. Therefore, you should include any tools and media that you normally sterilise in the autoclave.
  • If you will incubate the sample on site, be sure to swab the spore test fluid onto the agar plate in a sterile environment (e.g., a laminar flow hood).
  • The spore test contains bacteria that may pose a health hazard. Dispose of the test properly through sterilisation or incineration.
  • Do not continue to use an autoclave if it fails the spore test.

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