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Sterility Assurance in Dental Instrument Reprocessing

Biological indicators, when used in combination with other steam sterilization monitoring protocols, are an excellent way to ensure sterility during dental instrument reprocessing.

When used in combination with other steam sterilization monitoring protocols, Biological indicators are an excellent way to ensure sterility during dental instrument reprocessing

The views expressed are those of the authors and do not reflect the official views or policy of the U.S. Department of Defense or its components.

Sterility assurance is a cornerstone of a successful infection prevention program, and an effective sterilization protocol will help ensure the delivery of safe care. Once dental instruments become contaminated, reprocessing for subsequent use is a multistep procedure that involves cleaning and disinfection or sterilization. Effective sterilization cannot be accomplished if instruments are not first properly cleaned, as proficient cleaning is vital to successful reprocessing.1 Instruments having the greatest risk of disease transmission are categorized as critical items (i.e., those used to penetrate the patient’s bone or tissue) and must be heat sterilized. Instruments that touch mucous membranes have a lower risk of disease transmission and are categorized as semicritical items.2 The majority of semicritical dental instruments and dental slow- and high-speed handpieces are heat tolerant, and the U.S. Centers for Disease Control and Prevention (CDC) recommends these also be heat sterilized.3

The most popular steam sterilizers used in clinics are either gravity-displacement units that utilize gravity to remove air as it is displaced by steam, or dynamic air-removal devices (e.g., prevacuum sterilizers) that remove air by preconditioning techniques before steam and pressurization occur within the chamber.4 The biological indicator (BI) test, also known as the spore test, provides a direct assessment of the sterilizer’s lethality by killing highly resistant bacterial spores. This is the most widely accepted method for monitoring steam sterilization.5 However, if BI testing is the only monitoring method used for instrument reprocessing, it is possible to have nonsterile instruments even with passing BI test results because BIs only test one aspect of instrument reprocessing. So how can dental teams confirm instrument sterility?

Although BI testing is an important tool, it may not paint a complete picture of instrument sterility. Consequently, BIs should not be the only method used to verify sterilization. Sterility assurance level (SAL) is the probability that a single microbe would remain on an instrument after sterilization. SAL is often expressed as 10-n. Throughout the world, SAL for medical instruments is widely accepted as 10-6, which translates to a one in 1 million probability of finding a single living organism on medical instruments following sterilization.6 Nevertheless, if the bacterial spores within the BI are killed after sterilization, and the test thus yields passing results, this does not prove sterilization was satisfactory throughout the load — or even within each individual package of instruments.4,7 Besides BI monitoring, additional aspects of dental instrument reprocessing should be verified to assure sterility. For example, if instruments were not adequately cleaned before sterilization or if the sterilant (steam) did not reach the inside of packages for the required time and temperature — or if package integrity is compromised — nonsterile instruments could result, despite passing BI test results.


As noted, the first critical step to reprocessing is cleaning.4 Before instruments can be sterilized, they must be free of bioburden. Unless the steam sterilant directly contacts all surfaces, the instruments will not be sterile.8 Bioburden and debris can impede the sterilant from reaching the surface. If cement is discovered chairside on a dental instrument that has been reprocessed, removing the cement can expose a surface that steam did not reach and viable microorganisms could exist. For these reasons, it is imperative that the cleaning processes be effective and monitored. Carefully preclean instruments to remove gross contaminates during or immediately after the dental procedure. Using a pretreatment spray, gel or foam at the point of use is recommended to prevent blood, saliva and other bioburden from drying on instruments, which make them more difficult to clean.4

It is important to utilize cleaning methods consistent with the instrument manufacturer’s instructions for use (IFU). When a choice exists between manual and automated cleaning, choose the latter. Manual cleaning is less efficient than automated cleaning, time consuming, not consistently reproducible, and may lead to injury. Automated cleaners (such as ultrasonic cleaners and washer/disinfectors) provide reliability, reproducibility and can be validated.4,9 After the cleaning cycle, verify the instruments are clean through visual inspection. Magnification and lighting should be considered. Repeat the cleaning if contamination is found during visual inspection. Cleaning confirmation is also accomplished by conducting daily verification testing of cleaning equipment. This will determine if manual cleaning was accomplished properly and if automated equipment is operating optimally.4,10 It is imperative to recognize that instruments contaminated with visible debris should not be sterilized.


Once the cycle is complete, physical monitors of time, temperature and pressure provide a real-time evaluation of the sterilization process. Unlike the results of a BI test, this assessment is immediate. Results determining whether the load met the required time and temperature parameters of the manufacturer’s IFU are captured and documented by means of a printout or electronic format.4 The IFU for each instrument must be followed to ensure sterility;10 for example, items with lumens (e.g., dental handpieces) may require longer sterilization times to ensure steam reaches not only the surfaces, but the internal components, as well. Because sterilant only works if it reaches intended areas of instruments, it is important to know the manufacturer’s required time and temperature for sterilization and monitor the sterilization phase to ensure the intended parameters are met.

The temperature of the cycle is dictated by the manufacturer’s IFU, and some cycles may require higher temperatures than others. Manufacturer guidelines that require higher temperatures may have shorter cycle times, consequently, reviewing the IFU is essential to ensure the sterilizer is set appropriately. It is also important to complete physical monitoring immediately after the cycle and before BI results are available to establish real-time verification the intended parameters were met.4 In addition to time, temperature and pressure, other critical components should be verified after each sterilization cycle.

Chemical indicator (CI) monitoring verifies that one or more specific conditions of the sterilization cycle were achieved — either inside individual packages or at a specified location. Some CIs may do both. There are six types that vary based on the specific cycle characteristics they react to. Type 1 CIs, also known as external indicators, are most often placed on the outside of individual packages and help dental teams clearly differentiate packs that have been exposed to the sterilization process from those that have not. Type 2 CIs (e.g., Bowie Dick tests) are designed for use in very specific sterilization test procedures. The Bowie Dick test is only used in prevacuum sterilizers and determines if air is adequately removed from the chamber during the sterilization cycle.

Types 3, 4, 5 and 6 are internal CIs and designed to react to different critical sterilization variables at various stated values. Type 3 CIs react to a single variable, Type 4 indicators react to two or more variables, and Type 5 (also known as integrating indicators or integrators) react to all the critical variables of sterilization: temperature, time and pressure. According to the Association for the Advancement of Medical Instrumentation, the critical parameters for a passing Type 5 indicator are equivalent or exceed performance values of BIs (although this does not allow replacing BI testing with Type 5 CIs). Only a BI verifies the lethality of the sterilizer because a BI contains live spores. Instrument processing standards recommend BI testing at least weekly.4 Similar to Type 5 CIs, Type 6 indicators also react to all variables, but are designed for specified sterilization cycles.4

In addition to monitoring individual packages, Type 5 and Type 6 CIs can be used individually within a process challenge device to monitor an entire sterilization load. At least one internal CI should be placed in each package intended for sterilization.10 Internal CIs are particularly valuable because they can verify that steam reached the inside of the package and, subsequently, the instruments. Overpacking instruments in packages, overloading the sterilizer, poor orientation of packages, or air pockets trapped in the sterilizer can prevent steam from entering packages and result in CI failures.

As previously stated, BI monitoring verifies conditions where the BI test was located were sufficient to kill highly resistant bacterial spores. For steam sterilization, BI tests should contain spores of Geobacillus stearothermophilus. There are numerous types of BIs for steam sterilization, and each has unique characteristics — for example, for use in specified cycles (times and temperatures) and specific sterilizers (gravity displacement or dynamic air-removal units). Therefore, it is imperative to select the correct BI for a given sterilization cycle.4 Although the BI provides valuable information, the results are not available immediately after the sterilization cycle, as they may take up to 48 hours. For this reason, it is important to use a monitoring method that provides sterility assurance immediately after the cycle.


Package integrity should be verified after the sterilization process, before instruments are stored, and again immediately before opening the package for use.10 Once the sterilization cycle is complete and the physical monitors have been verified, dental instruments should be allowed to cool and dry. If packages contain moisture externally or internally after sterilization and appropriate cooling, they should be considered contaminated. In addition to moisture, packages should also be inspected for damage (e.g., tears, stains or improper seals) and to identify any external CIs and internal CIs (if visible) that did not change appropriately. If such packs are found, they should be reprocessed.4,10 Additionally, it is excellent practice to inspect the packages when they are taken from storage to the operatory. Finally, during operatory setup, the packages should again be inspected. Developing package integrity checks decreases the risk of using nonsterile instruments, even when loads pass BI testing.


Although transmission of infectious agents among patients and providers in dental settings is rare, it does occur. According to the CDC, these incidents have been linked to poor infection prevention procedures — including a lack of policies for steam sterilization of handpieces and failure to conduct sterilizer monitoring.2 When used in combination with other monitoring methods, BI testing is an excellent way to assure instrument sterility. Physical monitoring can help identify problems with the sterilizer equipment or issues with a single cycle. Similarly, CI monitoring can determine if there is a problem with improper packaging, loading or sterilizer malfunctions. In concert with these protocols, BIs can identify operator error or verify whether sterilizer conditions have the ability to kill spores. Finally, package integrity checks help determine whether the sterility of the package has been compromised following the sterilization process. Together, these methods will provide sterility assurance — and peace of mind — that the dental team has achieved successful sterilization of reprocessed instruments.


  • Reprocessing dental instruments is a multistep procedure that involves cleaning and disinfection or sterilization.
  • A biological indicator (BI) test, also known as the spore test, provides a direct assessment of the sterilizer’s lethality by killing highly resistant bacterial spores, but BI monitoring should not be the only method used to verify successful sterilization.
  • Physical monitors of time, temperature and pressure can help identify problems with sterilizer equipment or issues with a single cycle.
  • Similarly, chemical indicator monitoring can determine if there is a problem with improper packaging, loading or sterilizer malfunctions.
  • Package integrity should be verified after the sterilization process, before instruments are stored, and again immediately before opening the package for use.10


The authors wish to thank Helene Bednarsh, BS, RDH, MPH, for her diligent review and feedback.


  1. Whitworth CL, Martin MV, Gallagher M, Worthington HV. A comparison of decontamination methods used for dental burs. Br Dent J. 2004;197:635–640.
  2. U.S. Centers for Disease Control and Prevention. Summary of Infection Prevention Practices in Dental Settings: Basic Expectations for Safe Care. Available at: www.cdc.gov/oralhealth/infectioncontrol/guidelines/index.htm. Accessed July 11, 2018.
  3. Kohn WG, Collins AS, Cleveland JL, et al. Guidelines for infection control in dental health-care settings — 2003. MMWR Recomm Rep. 2003;52:1–61.
  4. Association for the Advancement of Medical Instrumentation. ANSI/AAMI ST79:2017. Available at: www.aami.org/productspublications/ProductDetail.aspx?ItemNumber=1383. Accessed July 11, 2018.
  5. Cuny E. The use of a process challenge device in dental office gravity displacement tabletop sterilizers. Am J Infec Control. 2015;43:1131–1133.
  6. van Doornmalen J, Kopinga K. Review of surface steam sterilization for validation purposes. Am J Infec Control. 2008;36:86–92.
  7. Smith AJ, Bagg J, Hurrell D, McHugh S. Sterilization of re-usable instruments in general dental practice. Br Dent J. 2007;203:E16.
  8. Dion M, Parker W. Steam sterilization principles. Available at: www.ispe.gr.jp/ISPE/07_public/pdf/201504_en.pdf. Accessed July 11, 2018.
  9. Walker JT, Dickinson J, Sutton JM, Raven ND, Marsh PD. Cleanability of dental instruments — implications of residual protein and risks from Creutzfeldt-Jakob disease. Br Dent J. 2007;203:395–401.
  10. Seavey R. High-level disinfection, sterilization, and antisepsis: current issues in reprocessing medical and surgical instruments. Am J Infec Control. 2013;41:S111–S117.

The authors have no commercial conflicts of interest to disclose.


From Decisions in Dentistry. August 2018;4(8):14,17–18.

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