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Autoclave Efficacy Validation for Biological Waste

The U of M has previously required that a cycle length for waste autoclaving is to be a minimum of 1 hr autoclave

time @121 C (40 minutes @132 C) unless you can prove valid decontamination in less time.

The Current Public Health Agency of Canadas Laboratory Biosafety Guidelines requirement for autoclave efficacy monitoring is found in Chapter 3.1.1 as follows. Chapter 8 also has a short discussion on autoclaves

3.1.1 18. Contaminated materials and equipment leaving the laboratory for servicing or disposal must be appropriately decontaminated and labeled or tagged-out as such.

3.1.1 19. Efficacy monitoring of autoclaves used for decontamination with biological indicators must be done regularly (i.e., consider weekly, depending on the frequency of use of the autoclave), and the records of these results and cycle logs (i.e., time, temperature and pressure) must be kept on file.

The following are resources for meeting these requirements: 1. Bags should not be packed more that full. 2. Bags should be loosely secured. Steam needs to penetrate in order to sterilize the load. 3. Validate autoclaving efficacy with standardized maximum loads that reflect actual use scenarios. These are

then the maximum size and distribution of bags/articles that are not to be exceeded in subsequent loads. 4. The Biological Indicator (BI) test vials should be placed in the worst-case scenario spots in the load. This is

the hardest place for steam to penetrate and the coldest part of the autoclave chamber during an autoclaving cycle. In a typical autoclave this is usually near the bottom and centre of an autoclave bag and toward the bottom front of the autoclave (approaching the drain).

5. You can facilitate retrieval of the vial after autoclaving by attaching a brightly coloured paper or a long string to the biological indicator vial before placing it in the worst-case spot in the load.

6. Because you must assume that the load may not be completely sterilized before you have information from the biological indicator testing, you may want to run a typical test load that is not biologically hazardous first. If not, as you retrieve the vial from the autoclaved bag, be sure you wear gloves and work carefully, surface disinfecting the outside of the vial as soon as you have removed it.

7. After being exposed to an autoclave cycle, the BIs are incubated with a culture medium to evaluate for a negative result. A negative result indicates no sign of bacterial growth/recovery (this is an observation of no turbidity and colour change). Vendors specify the incubation temperature (usually 56-60C) for a specified time frame (usually 24-48 hours) to evaluate for growth.

8. A control vial which has not been autoclaved must be incubated at the same time and temperature as the test vial. It must also be from the from the same lot# as the test vial. The Control vial should show a positive result. That is, after incubation it should show good growth of the bacterial culture as indicated by turbidity and colour change. This indicates that your batch of BI is active.

9. Biological Indicators are available in a variety of formats. These are discussed on page 2 along with a list of a few suppliers.

10. To determine the minimum autoclave cycle times based on biological indicator testing, consider doubling the minimum autoclaving time that is required for a test vial of the biological indicator to show a negative result.

11. A log book which logs all of the waste autoclaving cycles which are run should be maintained. A sample page with the recommended basic fields for a log book can be found at the end of this brief.

12. After establishing standardized loads and times as determined above, autoclaving efficacy should be validated on a regular basis with biological indicator testing in an as used scenario. For example, consider regular as:

a. Weekly if one or more waste autoclave loads are run daily b. Every two weeks if 2-3 loads/week c. Monthly if < one load/week.

13. The actual autoclave cycle recordings should be kept for three years (CFIA). Cycle recordings are usually circular charts or long paper roles.

14. If the autoclave does not have a cycle recorder to validate that the predetermined cycle parameters (time, temperature/pressure) have been achieved, then Chemical Sterilization Integrators must be used with every waste autoclaving load. Ordering resources for some typical chemical sterilization integrators are found below.

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15. It is also highly advisable that the department keeps a record of autoclave maintenance and repair and considers annual servicing of the autoclave that includes temperature verifications.

Autoclave Efficacy Validation Background Information

The parameters of 121C (15psi) for 12 minutes have been scientifically determined to kill highly resistant microorganisms in an autoclave. Cycle times of 20-30 minutes for wrapped goods are usually chosen to allow for chamber conditioning and a safety factor. A review of the literature however will also clearly indicate that standardization of waste autoclave parameters (time, temperature/pressure, load size, composition and distribution, steam quality) must be validated in an as used scenario. The composition and thickness of the autoclave bag and support container, as well as the cycle type, load distribution, and density, can all significantly increase the total cycle times needed to achieve adequate kill in the centre of a container, even when the chamber temperature pressure and time parameters have been met.

A variety of products from a number of vendors are available to assist in this process. Autoclave tape: Autoclave tape is a type of chemical indicator. It only indicates that the material to which it is attached has been processed. It must be used on all biological waste bags and sharps containers to distinguish autoclaved from non-autoclaved material. It does not confirm kill.

Biological Indicators: Biological Indicators (BIs) are used to determine if an autoclave has delivered a lethal cycle. Spores of Geobacillus stearothermophilus are usually used. These spores are very heat resistant. After being exposed to an autoclave cycle, the BIs are incubated with a culture medium to evaluate for growth/recovery (usually observation of turbidity and colour change). Vendors specify the incubation temperature (usually 56-60C) for a specified time frame (usually 24-48 hours) to evaluate for growth. Biological Indicators are available in a variety of formats. A description of some of the formats follows. The self-contained units would be the most commonly used for non-liquid waste loads. Always follow manufacturers recommendations for use. Spore Strips-spores inoculated onto filter paper and packaged in paper pouch or envelope. These require asceptic transfer to a growth media for incubation. The media must be made or purchased separately. Self-contained units- Here a sealed glass ampoule with recovery media is housed in a plastic tube along with the spore strip. After autoclaving and cooling, the cap on the plastic tube is shut and the glass vial is crushed, mixing the spores with the media for incubation. Most vendors have special incubators available for incubating their vials. A similarly crushed control vial that hasnt been autoclaved is incubated concurrently. Sealed ampoules- These have spores suspended in recovery media sealed inside a glass ampoule. No crushing or transfer is required. As steam condenses on the glass, the energy from the steam is transferred across the glass to the liquid media within the ampoule. (see reference #4). Because these are sealed units, they are appropriate to validate efficacy of sterilization in liquid media. Check with the vendor information for their use in non-liquid loads.

Sterilization integrators: Sterilization Integrators are chemical indicators that have been designed to provide accurate, immediate visual confirmation that one or more of the established autoclave sterilization cycle parameters (time, temperature, steam) have been met. No chemical indicator verifies that an item is actually sterile. These should be used in every waste autoclave load if you do not have an autoclave cycle recorder e.g. circular chart or paper role etc.

Examples of some efficacy monitoring products ( 2007 research): Biological Indicators: 3M Attest 1262-S Biological Indicator for steam Fisher Scientific carries this EM Science Sterikon Plus Bioindicator (Ampoule type) VWR carries this- EM1.10274.004 SGM BioTech EZTest Getinge Biosign Indicators Catalogue No.61301600022/21 50 per box/10 per box (a number of autoclave owners at the U of M use these.) Raven Labs ProSpore2 Steam, Prospore Ampoule

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Chemical Sterilization Integrators: Raven Labs ProChem EXT 3M Comply SteriGage Sterilization Integrator VWR carries this - 3M#1243A; VWR #34010-19 References: 1. Practical Guide to Autoclave Validation by Raymond G. Lewis,PE

http://www.ch2m-idcfs.com/Papers/IDC2002%20autoclave.pdf 2. Application Of Autoclaves In The Treatment Of Medical Waste

Poster Presentation Displayed at the 50th Annual Conference of the American Biological Safety Association October 7 10, 2007 Nashville, TN Ira F. Salkin1, Timothy Barrett2, Edward Krisiunas3, Shawn Doyle4, and Paul Warden5 1Information From Science, West Sand Lake, New York, 2OnSite Sterilization, Inc.,

http://www.ozonatorindustries.ca/resources/ABSAHandout.pdf 3. Chemistry of Indicators, SteriTec Products Inc.

http://www.steritecproducts.com/PDF/TBChemisrytofIndicators.pdf 4. Performance Comparison for Three Types of Biological Indicators Used in Steam Sterilization Processes:

Spore Strips, Crushable Self-contained and Sealed Glass Ampoule http://www.ravenlabs.com/pdf/asmpre.pdf

5. Steam Sterilization and BI Monitoring Liquid Loads/ Liquid Media

http://www.ravenlabs.com/pdf/SteamSterilizationAndBIMonitoring.pdf 6. 3M Attest 1262-S Biological Indicator for Steam: Product Profile http://multimedia.3m.com/mws/mediawebserver?66666UuZjcFSLXTtmxTyoxf6EVuQEcuZgVs6EVs6E666666-- 7. Study Guide#018, Steam Sterilization, A Continuing Education Study Guide presented by STERIS. 8. Favero, M.S. Developing indicators for monitoring sterilization. In: Rutala, W.A. Disinfection, sterilization

and antisepsis in health care. Washington, DC: Association for Professionals in Infection Control and Epidemiology Inc., 1998;119-32.

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Biological Indicator LOG AUTOCLAVE SERIAL # ________________________ AUTOCLAVE LOCATION ________________________

Cycle Date & Time

Cycle and Temperature Cycle Length Incubation reading Date and Time

Biological Indicator and Lot#

Result- Pass/Fail

Autoclaved Indicator

Control Indicator