The choice of sterilisation method is not always simply "autoclave everything." Many medical devices and pharmaceutical products are heat-sensitive and cannot withstand steam sterilisation temperatures. Others are moisture-sensitive. Some are too large for standard autoclave chambers. The right choice depends on the material composition, the geometry of the device, the intended application, regulatory requirements, and validation evidence.
This page covers the four principal sterilisation methods used in healthcare and pharmaceutical manufacturing: steam (moist heat), ethylene oxide (EO), radiation (gamma and electron beam), and low-temperature gas plasma (hydrogen peroxide plasma). For each method: the mechanism of action, the materials it is compatible with, the key process parameters, the validation approach, and the residual considerations.
Steam Sterilisation: The Gold Standard for Heat-Stable Items
Steam sterilisation (autoclaving) using saturated steam at 121 or 134 degrees Celsius is the preferred method for all heat-stable, moisture-tolerant materials: surgical instruments (metal), drapes, gowns, laboratory media, glass labware, and pharmaceutical bulk solutions. It is the simplest, most reliable, most cost-effective, and best-characterised sterilisation method. It leaves no toxic residuals. The F0 concept allows quantitative verification of lethality (see Topic 14).
Limitations: cannot be used for heat-sensitive materials (plastics that melt or deform above 100 degrees Celsius, most electronic components, lubricants that degrade with moisture), materials that absorb moisture and are damaged by it, or items with long, narrow lumens that steam cannot penetrate reliably.
Pre-vacuum (prevacuum) cycles are used for porous loads (wrapped surgical packs, gowns, drapes) to ensure steam penetration by vacuum-assisted air removal before steam admission. Gravity displacement cycles are used for liquids and unwrapped non-porous items.
Flash sterilisation: 134 degrees Celsius, 3 to 4 minutes exposure in a prevacuum cycle, used for immediately needed surgical instruments when reprocessing time is limited. Flash sterilisation is intended as an emergency measure for unpackaged items, not as a routine replacement for packaged terminal sterilisation.
Ethylene Oxide: The Low-Temperature Solution
Ethylene oxide (EO or EtO) is a highly effective alkylating agent that kills microorganisms by irreversibly alkylating nucleic acids and proteins. It is used for sterilisation of heat-sensitive medical devices, including complex multi-component devices with electronics, catheters, single-use endoscopes, and devices with long lumens.
EO sterilisation operates at 37 to 63 degrees Celsius, much lower than steam, and can penetrate most packaging materials and device lumens. The cycle consists of: pre-conditioning (warming and humidification of the load, typically at 40 to 60 per cent relative humidity, to ensure EO penetration), gas admission (EO gas alone or mixed with CO2 or nitrogen for safety), exposure (typically 1 to 6 hours depending on load and cycle parameters), and aeration (removal of EO and its toxic breakdown products from the load).
The critical safety concern: EO is a highly toxic, potentially carcinogenic gas. After sterilisation, significant EO and ethylene chlorohydrin (a toxic breakdown product) residuals remain in the device and its packaging. Aeration (at elevated temperature, typically 50 to 60 degrees Celsius, for 8 to 24 hours) reduces residuals to safe levels defined in ISO 10993-7. EO facilities require dedicated ventilation, atmospheric monitoring, and worker exposure management.
EO is increasingly subject to regulatory restrictions due to its environmental and occupational health impact. In some regions (including certain US states and EU member states), the air emissions from EO sterilisation facilities are under increasing regulatory pressure. Alternative low-temperature sterilisation methods are growing as a result.
Radiation Sterilisation: Gamma and Electron Beam
Radiation sterilisation uses ionising radiation (gamma rays from cobalt-60 sources, or electron beams from linear accelerators) to kill microorganisms by creating free radicals that damage DNA, cell membranes, and proteins irreversibly.
Gamma radiation: high penetrating power, can sterilise bulk packaged products (boxes, pallets) in a single pass through the irradiator. Used for single-use medical devices (syringes, surgical gloves, surgical sutures, implants, blood bags), pharmaceutical products, and tissue allografts. The standard sterilisation dose is 25 kGy (kilograys), though dose setting is validated for each specific product bioburden level using the ISO 11137 dose-setting methods.
Electron beam (e-beam) radiation: delivers a very high dose rate (much faster than gamma), but has lower penetration (typically less than 10 cm in density 1 material). Used for surface or low-density products where high-speed processing is an advantage.
Advantages: no toxic residuals, can penetrate sealed packaging and bulk stacks, no requirement for temperature or humidity control.
Limitations: some materials degrade under radiation (certain polymers, elastomers, electronic components): radiation compatibility must be validated before use. Products must be shipped to a central irradiation facility (not performed in-house except at very large manufacturers).
Low-Temperature Hydrogen Peroxide Gas Plasma
Hydrogen peroxide gas plasma sterilisation operates at temperatures below 55 degrees Celsius and uses a combination of hydrogen peroxide vapour and plasma (ionised hydrogen peroxide gas activated by radiofrequency energy) to kill microorganisms. The system kills by oxidation of cellular components through reactive oxygen species.
The primary clinical application is sterilisation of endoscopes and complex surgical instruments that cannot be steam-sterilised (heat-sensitive, moisture-sensitive) or EO-sterilised (do not tolerate long cycle times or need rapid turnaround). The STERRAD system (ASP, a Johnson and Johnson company) is the most widely used system globally.
Advantages: rapid cycles (28 to 75 minutes depending on the system), no toxic residuals, no aeration required, safe for electronics and heat-sensitive materials, low environmental impact.
Limitations: cannot sterilise liquids, cannot penetrate cellulosic materials (paper, cloth), requires special non-woven polypropylene or Tyvek packaging (not standard paper wrapping), cannot sterilise items with very long narrow lumens (hydrogen peroxide cannot reach the lumen end) without specific adapters.
The Sporicidal Hierarchy: Disinfection vs Sterilisation
It is important to distinguish sterilisation from disinfection, and high-level disinfection (HLD) from intermediate- and low-level disinfection.
High-level disinfection kills all microorganisms except high numbers of bacterial spores: used for heat-sensitive, semi-critical devices such as flexible endoscopes that cannot be steam-sterilised. Agents: glutaraldehyde (2 per cent, 20 to 45 minutes), ortho-phthalaldehyde (OPA, 0.55 per cent, 12 to 15 minutes), peracetic acid, hypochlorite at high concentration.
Sterilisation eliminates all microorganisms including spores to SAL 10^-6: the methods described on this page. High-level disinfection is not sterilisation, and the two should not be conflated in clinical practice.
Spaulding classification: critical items (penetrate sterile tissues or vascular system: surgical instruments, implants, needles) must be sterile. Semi-critical items (contact mucous membranes or non-intact skin: flexible endoscopes, respiratory therapy equipment) require HLD at minimum. Non-critical items (contact intact skin: blood pressure cuffs, stethoscopes) require intermediate or low-level disinfection.
Frequently Asked Questions
What is sterilisation?
Sterilisation is the process of eliminating all viable microorganisms, including bacterial spores, from a product, device, or surface. The internationally required standard is a sterility assurance level (SAL) of 10^-6: a maximum probability of one viable organism surviving per million items processed.
When should steam sterilisation be used?
Steam sterilisation should be used for all heat-stable (withstanding 121 to 134 degrees Celsius), moisture-tolerant materials. It is the preferred method for metal surgical instruments, glass labware, gowns and drapes, and pharmaceutical aqueous solutions. It leaves no residuals, is well-characterised, and is the most reliable method for compatible loads.
Why can't endoscopes be steam-sterilised?
Modern flexible endoscopes contain complex optical fibres, electronic components, adhesives, and heat-sensitive polymers that cannot withstand autoclave temperatures. They also contain long, narrow internal channels (for air/water and suction) that steam cannot reliably penetrate. They require high-level disinfection with chemical agents or sterilisation using low-temperature methods (hydrogen peroxide gas plasma or EO).
What is ethylene oxide sterilisation used for?
EO sterilisation is used for heat-sensitive, single-use medical devices including complex multi-component devices with electronics, catheters, single-use surgical kits, and devices with narrow lumens. It is the most widely used alternative to steam sterilisation for medical devices globally, though it is under increasing regulatory scrutiny due to its toxicity and environmental impact.
What dose of radiation is used for sterilisation?
The standard radiation sterilisation dose for medical devices is 25 kGy (kilograys), though this can be adjusted up or down based on the validated bioburden of the specific product using ISO 11137 dose-setting methodology. A minimum dose of 25 kGy is sufficient to achieve SAL 10^-6 for products with a bioburden of up to 10^6 organisms per item, assuming a worst-case D-value for the reference organism.
What is the Spaulding classification?
The Spaulding classification (1968, E.H. Spaulding) categorises medical devices by their risk of infection transmission and the required level of reprocessing. Critical items (penetrating sterile tissues) must be sterile. Semi-critical items (contacting mucous membranes) require high-level disinfection at minimum. Non-critical items (contacting intact skin) require intermediate or low-level disinfection.
What is high-level disinfection?
HLD kills all microorganisms except large numbers of bacterial spores. It is the required minimum standard for flexible endoscopes and other semi-critical devices. Agents include 2 per cent glutaraldehyde, 0.55 per cent OPA (ortho-phthalaldehyde), peracetic acid, and high-concentration hydrogen peroxide. HLD is not sterilisation and should not be applied to critical devices.
What are EO residuals and why do they matter?
After EO sterilisation, residual EO and its breakdown product ethylene chlorohydrin remain in the device and packaging. Both are toxic: EO is a mutagen and potential human carcinogen (IARC Group 1). Aeration after the sterilisation cycle reduces residuals to levels defined as safe by ISO 10993-7. Devices must not be released for clinical use until aeration is complete and residuals are verified within safe limits.
What is the difference between gamma radiation and e-beam sterilisation?
Gamma radiation from cobalt-60 sources has very high penetrating power and can sterilise bulk packaged products. E-beam delivers a very high dose rate through a linear accelerator but has much lower penetration. Gamma is more versatile for bulk processing; e-beam is faster and more suitable for surface or low-density products where high throughput is needed.
Why is hydrogen peroxide plasma not suitable for all devices?
Hydrogen peroxide gas plasma cannot sterilise liquids (the water-absorbing) or cellulosic materials (paper, cloth: these absorb and inactivate the hydrogen peroxide before it can sterilise). Items with very long or very narrow lumens may not achieve adequate hydrogen peroxide concentration at the lumen tip without specific lumen adapters. Packaging must be non-cellulosic (non-woven polypropylene or Tyvek).