How long should an autoclave hold at 121°C? For many laboratory loads, a holding time of 15–20 minutes after the load reaches sterilization
temperature is commonly used. However, the appropriate holding time depends on the materials, load size, liquid volume, container type, and
how the autoclave is loaded.
This guide explains what autoclave holding time means, why it varies, and how monitoring the temperature of the load can support more reliable
sterilization.
What is autoclave holding time?
Autoclave holding time, sometimes called exposure time, is the period during which the materials being sterilized remain at the required sterilization temperature.
Holding time is not necessarily the same as the total cycle time. A complete autoclave cycle may include heating, air removal, sterilization,
exhaust, and cooling. The chamber may also reach the selected temperature before the center of the load does.
1
Heating begins
The autoclave chamber and load begin warming.
2
Chamber reaches temperature
The chamber may reach the target before the load.
3
Load reaches temperature
Heat penetrates the materials being sterilized.
4
Holding time begins
The load remains at temperature for the validated duration.
Recommended autoclave holding times
The WHO Laboratory Biosafety Manual, 3rd Edition provides the following examples of steam sterilization cycles for correctly loaded autoclaves.
| Sterilization temperature | Minimum holding time |
|---|---|
| 121°C | 15 minutes |
| 126°C | 10 minutes |
| 134°C | 3 minutes |
These times are general examples, not universal cycle recommendations.
Load configuration, material type, volume, and applicable procedures must also be considered.
All materials to be autoclaved should be placed in containers that allow air removal and heat penetration. The chamber should also be loosely packed so steam can reach the load evenly.
Adapted from the WHO Laboratory Biosafety Manual
Why autoclave holding times vary
There is no single holding time that is appropriate for every laboratory load. Several factors influence how quickly steam and heat reach the materials being sterilized.
Load size and density
Large or tightly packed loads can restrict steam circulation and require more time for uniform heating.
Liquid volume
Larger volumes take longer to reach the sterilization temperature than small bottles or partially filled containers.
Container shape
Tall, narrow, sealed, or densely filled containers can affect heat transfer and steam penetration.
Type of material
Glassware, media, instruments, liquids, and biological waste may require different cycle parameters.
Trapped air
Air pockets can prevent steam from contacting the load and reduce the efficiency of heat transfer.
Chamber loading
Overpacking the chamber or blocking steam pathways can lead to uneven heating throughout the load.
Why microbiological waste may require more time
The Centers for Disease Control and Prevention emphasizes that cycle conditions should be selected according to the individual load.
As one example, the CDC notes that a 10-pound load of microbiological waste may require at least 45 minutes at 121°C. Waste bags and densely packed materials may contain trapped air, which slows steam penetration and delays uniform heating.
Do not assume that the center of a waste bag or large liquid container has reached sterilization temperature simply because the chamber display reads 121°C.
Why measuring load temperature matters
An autoclave normally monitors the temperature inside the chamber.
However, the materials within the load may heat more slowly than the surrounding chamber, particularly when sterilizing large liquid volumes or densely packed materials.
The TOMY External Temperature Sensor Probe allows SX-Series users to monitor and display the temperature of the material being sterilized. This can help laboratories determine when the load has reached the selected temperature instead of relying only on chamber temperature.
Monitoring the load temperature also helps address heat-transfer time lag, the delay between the chamber reaching temperature and the load reaching the same temperature.
Autoclave holding time FAQs
Does holding time begin when the chamber reaches 121°C?
Not necessarily. The chamber may reach 121°C before the materials inside the load. Holding time should be based on the validated
sterilization procedure and the temperature reached by the load.
How long should liquids be autoclaved?
The required time depends on the volume, container, fill level, and loading arrangement. Large liquid volumes generally take longer to heat than small volumes and should be validated separately.
Why does laboratory waste require longer cycles?
Waste bags and densely packed loads may retain air and limit steam movement. This slows heat penetration, particularly near the center of the load.
Can an autoclave chamber be packed tightly?
No. Materials should be arranged so that air can escape and steam can circulate around the load. Overpacking can cause uneven heating and incomplete sterilization.
How can an autoclave cycle be validated?
Validation may include physical monitoring, load-temperature measurement, chemical indicators, biological indicators, and documented procedures appropriate for the application.
TOMY SX-Series
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