Autoclaves

Safe and Effective Use

Environmental Health and Safety

University of Central Florida

Topics

Sterilization Principles
 Safety Instruction

Proper Use of Cycles

Special Situations
 Autoclave sterilization-techniques

Safety concerns

Research concerns, and

UCF Biohazard Waste Policy
Principles of Steam Sterilization

Sterilization: the complete destruction of
all forms of microbial life, including
bacterial spores


Steam under pressure reaches high
temperatures
Principles of Steam Sterilization (cont.)

 The “killing power” of steam is due to its

latent heat of vaporization

80 calories: 1 L water to boiling

540 calories: 1 L boiling water to steam
 Latent Heat of Vaporization

For example:
Action of Steam

Steam contacts a cooler surface, condenses,
causing a huge decrease in volume and
setting up a negative pressure that draws
more steam

Condensation occurs as long as there is a
temperature differential

Action of steam ensures: Surface heating,
penetration, and protein coagulation
Thermal Death Time (TDT)

TDT is the time required to kill a known
population of microorganisms in a specific
suspension at a particular temperature


Increasing temperature decreases TDT


Lowering the temperature increases TDT
Thermal Death Time ( cont.)

Acidic or basic pHs decrease TDT


Fats and oils slow penetration and
increase TDT
Standard Temperature and Pressure


250 °F (121 °C) and 15 p.s.i.
Safety Instructions
 PPE - heat proof

Gloves

Face Shield

Apron
Safety Instructions (cont.)
 Proper procedures

Close door tightly

Check chamber pressure before
opening

Crack door before opening

Wear loose fitting thermal gloves

Let hot liquids cool
Hazard Warnings

 Explosion hazard: Autoclaves are

not designed to process flammable
liquids
Burn Hazard When Sterilizing Liquids

Before loading containers of liquids into
the autoclave, the caps must be loosened to
avoid having the bottles shatter during
pressurization.

Use vented closures on bottles.

Use Type 1 borosilicate glass bottles.

Avoid sudden full opening of door at the
end of the cycle. Crack one inch, wait 10
minutes.
Burn Hazard When Sterilizing
Liquids (cont.)

Use LIQUID cycle only.

Do not jolt hot bottles. Do not move
bottles if bubbling or boiling.

Allow bottles to cool to touch before
attempting to move them from the
sterilizer shelf.
Hazard Warnings

 Slip hazard: Wipe up spills on slippery

floors.
 Burn hazard: Remember sterilizer racks
and shelves will be HOT after cycle is
run. Wear protective gloves, apron, and
face shield if needed.
Hazard Warnings (cont.)
 When removing items from the
autoclave, always wear a rubber apron in
addition to rubber sleeve protectors, heat
resistant gloves and a face shield.
 Remove the load and let the glassware
cool for 15 minutes before touching it
with ungloved hands.
Hazard Warnings

Allow sterilizer and everything else to cool
before cleaning or performing
maintenance.
 Burn and shock hazard: Repairs and
adjustments should be performed by
authorized personnel only.

Never clean sterilizer with abrasives, wire
brush, or steel wool.
Vocabulary
 Gravity

jacket is steam reservoir for chamber
 Isothermal (“same temperature”)

jacket and chamber charge with steam at the
same time and discharge at the same time
 Pre-vacuum

air is removed from the chamber before
steam is allowed in.
Sterilization Techniques
 Fabrics and Hard Goods

use gravity cycle (or pre-vacuum if
available)
 Liquid

use liquids cycle only
Fabrics and Hard Goods
(Gravity Cycle)
 Wrap in “blue surgical wrap”(available
from Boundary or Health Center Stores)

 Limit size and density

 Arrange load for maximum exposure

When Mixing Loads
 hard goods on lower shelves

 fabrics on higher shelves

 Sterilize liquids separately!

Liquids (Liquid Cycle Only)
 Use vented closures - no screw caps or rubber
stoppers
 Borosilicate glass (Pyrex) only
 Liquids cycle only
 Containers should be no more than two-thirds full
 When done, crack door 1/2 inch, wait 10 minutes
 Do not jolt hot bottles
 Move out cool bottles only
Operating Instructions
 Checklist before operating
 Automatic Operation

Gravity

Liquids

Laboratory/Isothermal
 Checklist Prior to Operation of
Autoclave

 Check settings
 Choose Gravity or Isothermal
 Check date/time
 Check paper tape roll
 Check printer cartridge
How Long to Autoclave (1)
Dry goods @ 121 °C Time (min)

 Glassware, empty, inverted 15

 Instruments, wrapped 30

 Utensils, wrapped 30
How Long to Autoclave (2)
Biohazardous waste bags, @ 121 °C, loosely tied
Time (min)

2 or more bags 100 +

1 bag, full 90

1 bag, half full 60

Testing available for individual situations

How Long to Autoclave (3)
Liquids, in bottles with vented caps
Size (ml) Time (minutes)
75 25
250 30
500 40
1000 45
1500 50
2000 55
Special Situations
 Media boils out of bottles

differs among media
 Very dense or closed dry objects

beaker full of microfuge tubes with
aluminum foil covering

SPF rodent cages
Summary

 We have gone over sterilization, safety, use

of cycles, and special cases

 Users should feel more confident about

their safety as well as their ability to assess
their loads and to choose the proper cycle
Where to get more information
 EH&S, Biological Safety

José Vázquez - 823-6301