Decontamination in healthcare and industry follows three progressively aggressive levels: cleaning, disinfection, and sterilization. Each level destroys a broader range of microorganisms than the one before it, and the choice of which level to apply depends on the risk an object poses to the person who will come into contact with it. This framework, first proposed in the late 1950s, still governs how hospitals, dental offices, food processors, and regulatory agencies think about making objects safe for use.
Why Three Levels Exist
The tiered approach traces back to the Spaulding classification system, originally put forward in 1957 by Earle Spaulding. The idea is straightforward: not every surface or instrument carries the same infection risk, so not every surface needs the same treatment. A bedside table that only touches intact skin is a very different proposition from a scalpel that enters sterile tissue. Spaulding sorted medical devices into three risk categories and matched each one to a minimum level of decontamination. Despite decades of advances in microbiology and infection control, this framework remains in wide use because it is logical, easy to apply, and understood by everyone from microbiologists to device manufacturers to regulators.1PubMed Central. A review of Spaulding’s classification system for effective cleaning, disinfection and sterilization of reusable medical devices
The three device categories, and their corresponding decontamination levels, work like this:
- Noncritical items: Objects that touch only intact skin, such as blood pressure cuffs and stethoscopes. These require low-level disinfection.
- Semi-critical items: Objects that contact mucous membranes or non-intact skin, such as respiratory equipment and flexible endoscopes. These require at least high-level disinfection.
- Critical items: Objects that enter sterile tissue or the bloodstream, such as surgical instruments and implants. These must be sterilized.
Cleaning sits underneath all of these as a prerequisite step. You always clean before you disinfect or sterilize, because organic matter left on a surface shields microorganisms from the chemicals or heat that are supposed to kill them.2PubMed Central. Disinfection and Sterilization in Health Care Facilities: An Overview and Current Issues
Level One: Cleaning
Cleaning is the physical removal of visible soil, organic matter, and a large share of microorganisms from a surface. It does not aim to kill every germ present. Instead, it reduces the microbial load enough that the next step, whether that is disinfection or sterilization, can work effectively. Think of it as preparing the battlefield rather than winning the war.
The tools of cleaning are water, mechanical action (scrubbing, flushing, or ultrasonic vibration), and detergents. Some facilities use enzymatic detergents that break down proteins and fats clinging to instruments, while others use detergents that contain low concentrations of germicidal agents like hydrogen peroxide. These combination products help protect healthcare workers from infectious risk during the soaking and scrubbing stages and reduce the number of organisms on a device before sterilization or disinfection takes over.3American Journal of Infection Control. A new hydrogen peroxide–based medical-device detergent with germicidal properties: Comparison with enzymatic cleaners
Why does cleaning matter so much as a first step? Because organic debris, blood, and mucus can physically coat microorganisms, preventing disinfectants and sterilants from reaching them. This is especially important for devices with narrow channels, like the tubing inside flexible endoscopes. Research has shown that washing endoscopes under high flow rates with effective detergents not only removes established biofilm but also retards new biofilm from forming, which underscores why thorough cleaning before disinfection is non-negotiable.4PubMed. The effect of multiple cycles of contamination, detergent washing, and disinfection on the development of biofilm in endoscope tubing
The Biofilm Problem
Biofilms deserve special attention because they represent one of the biggest challenges to effective decontamination at every level. When bacteria attach to a surface, they can form a sticky, self-produced matrix that shields the colony from chemical and physical attack. Cells deep inside a biofilm have reduced metabolic activity compared to free-floating cells, which makes them less susceptible to antimicrobial agents that target active cellular processes. For a disinfectant or sterilant to work, it first has to penetrate this matrix to reach the cells inside.5American Journal of Infection Control. 3 Levels of Decontamination: From Cleaning to Sterilization
This is precisely why cleaning is the essential first step. Mechanical scrubbing and high-flow flushing physically disrupt and remove biofilm, giving downstream disinfectants a much better chance of contacting and killing whatever organisms remain. Skip the cleaning step and you may end up with a device that has been soaked in a powerful chemical but still harbors viable organisms tucked safely inside a biofilm layer.
Level Two: Disinfection
Disinfection uses chemical agents (and sometimes physical methods like ultraviolet light) to kill most pathogenic microorganisms on a surface or device. It sits between cleaning and sterilization on the decontamination ladder, and it comes in its own sub-tiers: low-level, intermediate-level, and high-level disinfection. The difference between these tiers is defined by which organisms they can kill.
Low-level disinfection kills most vegetative bacteria, some fungi, and some enveloped viruses. It is appropriate for noncritical items that only touch intact skin. You encounter low-level disinfection constantly outside of hospitals: wiping down a gym machine, sanitizing a kitchen counter, or cleaning a phone screen with an alcohol wipe. In a clinical setting, wiping a stethoscope with an alcohol-based pad is a familiar example.
Intermediate-level disinfection adds the ability to kill mycobacteria (the organisms behind tuberculosis), most viruses, and most fungi. It does not reliably destroy bacterial spores. Hospital-grade disinfectants containing phenolics or specific concentrations of chlorine-based agents often fall into this category.
High-level disinfection kills everything except large numbers of bacterial spores. It is the minimum standard for semi-critical devices like endoscopes, laryngoscope blades, and respiratory therapy equipment. Common high-level disinfectants include glutaraldehyde, ortho-phthalaldehyde, hydrogen peroxide solutions at elevated concentrations, and peracetic acid. These chemicals achieve their killing power through mechanisms such as dissolving the lipid components of microbial cell membranes and denaturing the enzymes that microbes depend on for nutrient transport.6PubMed Central. Mechanisms of action of microbicides commonly used in infection prevention and control
When Disinfection Falls Short
The gap between high-level disinfection and sterilization is not just academic. Inadequate reprocessing of flexible endoscopes has been linked to outbreaks of healthcare-associated infections, including those caused by carbapenem-resistant bacteria, which are among the most dangerous drug-resistant organisms circulating in hospitals.7PubMed Central. Superbugs on Duodenoscopes: the Challenge of Cleaning and Disinfection of Reusable Devices These outbreaks have pushed some experts to argue that certain high-risk endoscopes, particularly duodenoscopes, should be reclassified from semi-critical to critical devices. That reclassification would mean they would need to be sterilized rather than merely high-level disinfected, increasing the margin of safety.1PubMed Central. A review of Spaulding’s classification system for effective cleaning, disinfection and sterilization of reusable medical devices
Level Three: Sterilization
Sterilization is the most aggressive level of decontamination. Its goal is the complete elimination or destruction of all forms of microbial life, including the tough bacterial spores that survive high-level disinfection. Any device that will enter sterile body tissue or the bloodstream must be sterilized before use.
The most common method is steam sterilization, often called autoclaving. Pressurized steam at high temperatures (typically around 121°C for a standard cycle, or 132–134°C for a faster cycle) denatures proteins and destroys microbial DNA. Steam sterilization is fast, reliable, nontoxic, and inexpensive compared to alternatives. It remains the gold standard for instruments that can tolerate heat and moisture, which includes most metal surgical instruments.
The challenge arises with devices that cannot withstand high heat and steam. Flexible endoscopes with delicate fiber optics, battery-powered instruments, plastics, and electronics all risk damage in a steam autoclave. For these items, healthcare facilities turn to low-temperature chemical sterilization methods.
Low-Temperature Alternatives
Ethylene oxide gas was the first chemical sterilant widely used in healthcare, adopted in the 1950s. It penetrates well into complex device channels and packaging materials, making it effective for items with lumens and crevices. However, ethylene oxide is toxic, potentially carcinogenic, and subject to stringent health, safety, and environmental regulations. Processing times are long, often requiring an extended aeration phase to allow residual gas to dissipate from the sterilized items. Because of these drawbacks, safer and more expedient alternatives have been introduced over the decades, including hydrogen peroxide gas plasma, vaporized hydrogen peroxide, and peracetic acid systems.8PubMed Central. Ethylene Oxide and Hydrogen Peroxide Gas Plasma Sterilization: Precautionary Practices in U.S. Hospitals
Hydrogen peroxide gas plasma has become one of the most popular low-temperature options. It works at temperatures below 55°C and has a relatively short cycle time, usually under an hour for many device loads. The process generates no toxic residues; the byproducts are water and oxygen. Its main limitation is that it does not penetrate very long or very narrow lumens as reliably as ethylene oxide, so it is not suitable for every device.
The Prion Exception
Prions are a category of infectious agent that breaks the rules of the entire decontamination hierarchy. Unlike bacteria, viruses, and fungi, prions are misfolded proteins with no nucleic acid, which means they have no DNA or RNA to destroy. This makes them astonishingly resistant to methods that work against every other pathogen. Standard autoclaving at 121°C, alcohol treatment, and even gamma-ray irradiation, which are reliable against bacteria and viruses, are not effective against prions.9International Journal of Molecular Medicine. Fundamentals of prions and their inactivation
Even more unsettling, prion infectivity can survive autoclaving at temperatures well above the standard range. Research has found that under certain conditions, the effectiveness of autoclaving actually declines as temperature is increased beyond 132–138°C. Strong sodium hypochlorite solutions (concentrated bleach) can achieve inactivation, though other chlorine-releasing compounds are less reliable. Sodium hydroxide at high concentrations leads to substantial but incomplete inactivation. Proprietary phenolic disinfectants perform even worse.10PubMed. Inactivation of prions by physical and chemical means
Because no routine reprocessing method guarantees complete prion inactivation, many guidelines recommend using single-use disposable instruments for procedures on patients known or suspected to have prion diseases like Creutzfeldt-Jakob disease. When reusable instruments must be used, protocols typically involve extended autoclaving at 134°C combined with sodium hydroxide immersion, though even these combined approaches leave some residual uncertainty. Prions represent the clearest example of a situation where the three-level decontamination framework reaches its limits.
Material Compatibility and Device Longevity
Choosing the right decontamination method is not only about killing microbes. It also has to account for what the process does to the device itself. High temperatures warp plastics. Harsh chemicals corrode metal coatings. Repeated cycling through cleaning, disinfection, and sterilization can degrade a device over its useful life, eventually compromising its function or safety.
Regulatory bodies have recognized this concern. ISO standards, EU medical device regulations, and FDA guidance all require manufacturers to determine whether repeated processing causes degradation that limits how many times a device can be safely reused. Testing protocols simulate dozens or even hundreds of reprocessing cycles, including soiling, cleaning, and sterilization, to measure cumulative wear. In one such assessment using a simulated 100-cycle protocol, researchers found negligible to nondetectable residual contaminants on both test coupons and actual devices, suggesting that well-designed instruments can hold up to heavy reprocessing loads when protocols are followed correctly.11PubMed Central. Assessing the Impact of End-of-Life Processing on Reusable Medical Devices
In practice, the material compatibility question is one reason why hospitals maintain an inventory of different sterilization technologies. An instrument that cannot go through steam autoclaving needs access to a hydrogen peroxide gas plasma or ethylene oxide system. This creates logistical complexity: sterile processing departments must track which devices are compatible with which methods, manage different cycle times and turnaround schedules, and ensure staff are trained on multiple systems.
Worker Safety Around Chemical Sterilants
The chemicals that make sterilization and high-level disinfection possible are, by definition, hazardous. If they were not potent enough to destroy microbial life in all its forms, they would not be useful. Glutaraldehyde, ethylene oxide, and peracetic acid can all cause respiratory irritation, skin sensitization, or more serious health effects with repeated exposure. Modern sterilizers are engineered to contain these chemicals safely, but mechanical failures, wear and tear, and user error can all lead to leaks.12PubMed. Safe use of chemicals for sterilization in healthcare
Workplace safety regulations require employers to assess the hazards of the chemicals in use, monitor for potential leaks, and ensure that worker exposure stays within permissible limits. For ethylene oxide, these regulatory requirements are especially stringent, which is one of the factors driving the shift toward hydrogen peroxide-based systems that produce only water and oxygen as byproducts. In facilities that still rely on ethylene oxide, ventilation systems, gas monitoring equipment, and strict aeration protocols are essential safeguards.
Beyond the Hospital
While the Spaulding classification was developed for medical devices, the same three-level logic applies across many industries. Food processing, pharmaceutical manufacturing, and laboratory science all rely on graduated decontamination strategies calibrated to the risk involved.
In the food industry, surface hygiene on production lines is critical for preventing contamination by pathogens like Salmonella and Listeria. The dry food industry faces a particular challenge because many traditional disinfection methods rely on water, which is incompatible with products like powdered milk, spices, and cereal. This has driven interest in dry disinfection technologies, including superheated steam, fumigation, alcohol-based disinfectants, ultraviolet radiation, and cold plasma.13PubMed Central. Process Technologies for Disinfection of Food-Contact Surfaces in the Dry Food Industry: A Review The underlying principle is the same as in healthcare: match the method to the risk and the material, and always start with thorough cleaning.
Cold atmospheric plasma is one of the newer technologies gaining traction across both healthcare and food safety. Plasma, generated at room temperature, produces a cocktail of reactive oxygen and nitrogen species that can inactivate bacteria, fungi, and viruses on contact while having minimal impact on heat-sensitive surfaces or food products.14PubMed. Integration of Cold Atmospheric Plasma and Light-Based Technologies for Surface Decontamination in Nonthermal Food Safety Applications Research on plasma-based decontamination in medical contexts has similarly shown it can efficiently destroy a range of pathogens, including on the surfaces of medical and dental devices.15PubMed Central. Disinfection and Sterilization Using Plasma Technology: Fundamentals and Future Perspectives for Biological Applications Whether plasma technologies will eventually displace established methods like autoclaving or hydrogen peroxide gas plasma in routine clinical use remains to be seen, but they represent a promising direction, especially for items and surfaces where traditional heat or chemical methods cause damage.
How Antisepsis Got Its Start
The entire enterprise of systematic decontamination in medicine is remarkably recent. Before the 1860s, surgeons operated with bare, unwashed hands, and instruments were reused without any attempt at cleaning between patients. Postoperative wound infections, gangrene, and death were considered inevitable consequences of surgery rather than preventable outcomes.
Joseph Lister changed that by connecting Louis Pasteur’s germ theory of fermentation to the problem of wound putrefaction. Lister promoted the use of carbolic acid (phenol) as an antiseptic during surgery, applying it to wounds, instruments, and dressings. The results were dramatic: wound sepsis and gangrene dropped sharply, which in turn reduced the need for amputations. By demonstrating that germs could be kept out of surgical wounds, Lister laid the groundwork for every subsequent advance in infection prevention.16PubMed Central. Joseph Lister (1827-1912): A Pioneer of Antiseptic Surgery
From Lister’s carbolic acid spray to today’s hydrogen peroxide gas plasma systems, the progression reflects a growing understanding of microbial diversity and resistance. Each time a new class of pathogen proved able to survive existing methods (bacterial spores in the early twentieth century, prions in the late twentieth century), the decontamination toolkit expanded. The three-level framework endures not because it is perfect, but because it forces the right question at the start of every reprocessing decision: how much risk does this object carry, and what is the minimum level of treatment needed to make it safe?