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Micro-08105 Practical 3
PRINCIPLE AND WORKING OF AUTOCLAVE
Dr. Shahzad Ali
Assistant Professor
Department of Wildlife and Ecology
UVAS, Ravi Campus, Pattoki
Autoclave
An autoclave is a pressure chamber used to sterilize equipment
and supplies by subjecting them to high pressure (15 psi)
saturated steam at 121 °C for around 15–20 (or more) minutes
depending on the size of the load and the contents
Uses of Autoclave
Surgical Instruments Plastic Sharps containers Glassware Plastic tubes and pipette tips Solutions and water Animal food and bedding Biohazardous waste
Principles of Autoclave Operation
• Steam penetrates objects in the autoclave
• Condensation creates negative pressure and draws in additional steam
• Moist heat kills microorganisms via coagulation of proteins
• Two types of autoclaves• Gravity Displacement
• Vacuum/Gravity Assisted
Types of Autoclaves
Other Manufacturers:
NOTE: Even though autoclaves on UNC campus operate similarly in the context of mechanics, training is required in the use of each autoclave since not all autoclaves are programmed the same.
Steris Castle Getinge
Preparation and Loading of Materials
Prepared the material for autoclaving with indicator paper/tape Fill he bottom of autoclave with water, up to support Place the material to be sterilized in the autoclave Always put bags of biological waste into pans to catch spills. Position biohazard bags on their sides, with the bag neck taped
loosely. Leave space between items to allow steam circulation. Household dishpans melt in the autoclave. Use autoclavable
polypropylene or stainless steel pans Close the lid and make sure that the rubber washer is in the
groove Screw down the clamps firmly Open the air outlet valve
Preparation and Loading of Materials Turn on the heating. Do not leave autoclave un-attended Close the outlet valve when a constant jet of steam issue from it Once the expected temperature is reached, reduce the heat to
maintain temperature Do not touch the drainage tape or the outlets or safety valves
while heating under pressure When the required time is up, turn off the heating completely When temperature falls below 100 ᵒC, open the outlet valve
slowly. Do not leave outlet valve opened for too long Never unscrew the lid clamps and open the lid until the hissing
sound has stop Leave the sterilized material to cool before removing it from
autoclave Check whether he autoclave tape (used as indicator) has turn
black
Indicator Tape (Heat Sensitive Tape):
Indicator tape in an “X” pattern.
Not autoclaved (no markings on tape)
Autoclaved (markings on tape)
Precautions for Time Selection
Take into account the size of the articles to be autoclaved. A 2-liter flask containing 1 liter of liquid takes longer to sterilize than four 500 mL flasks each containing 250 mL of liquid.
Material with a high insulating capacity (animal bedding, high sided polypropylene containers) increases the time needed for the load to reach sterilizing temperatures.
Autoclave bags containing biological waste should be autoclaved for 1 hour to assure decontamination.
In case of culture media mostly autoclaved at high pressure (15 psi) saturated steam at 121 °C (or higher) for around 15–20 minutes
Material Temperature Time
Laundry 121°C (250° F) 30 Minutes
Trash (Biohazard bags containing infectious waste.)
121°C (250° F) 1 Hour
Glassware 121°C (250° F) 1 Hour
Liquids 121°C (250° F) 15-20 Minutes
Animals 121°C (250° F) 8 Hours
Autoclave Waste Decontamination
Procedures
Precautions for Removing the Load
Put on Personal Protective Equipment Allow the autoclave to completely finish cycle
Pressure gauge must read zero Verify cycle conditions were met Open door slightly to allow steam to escape While slowly opening the autoclave door, make sure to
keep head, face, and hands away from the opening. Verify that heat sensitive tape has changed color or
word “autoclaved” has appeared. Allow contents to cool before removal When removing biohazard bags, always pick up from
the top, taped area of the bag. Never handle biohazard bags by grabbing from the sides or bottom.