Koch Lab:Protocols/How to Autoclave
The autoclave is one of the most useful tools in our lab. Without it much of our glassware and liquids will not be properly cleaned, leading to contamination, damage and worse of all, bad data. When you use the autoclave you want to ensure that you understand fully what you are doing to get the cleanest glassware possible. This page will guide you through the steps necessary to clean and dry the glassware and to sterilize liquids.
The Autoclave Controls
The controls for the autoclave will be mentioned several times in the steps below. There is one guage, a switch, three lights and three knobs. For a better understanding of how each of these components are related to each other, click on the thumbnail to the right for a larger photo. Roughly from top to bottom, each of the components are:
- Green start switch. This one is simple, flip the switch to start when using the autoclave and stop when not using it.
- Pressure gauge. This gauge is on the top left. The white hand measures the difference in pressure in psi between the inside and outside of the autoclave.
- Timer Knob. The top knob out of the three sets how long the timer will run in minutes. When the timer runs down to zero, it will ding to indicate that it is done.
- Temperature knob. The middle knob out of the three measures the temperature of the autoclave during the cleaning cycle. It is measured in both °C and °F. Centigrade is on the inside of the two numbers.
- Cycle knob. The bottom knob, which is separated from the other two, determines which part of the sterilizing cycle the autoclave is in. When you move this knob, always go clockwise. Clockwise from the top, the positions and functions of the autoclave are:
- "0" This is the default setting where the autoclave is doing nothing in particular. Whenever you are not cleaning something, and the autoclave is off, the knob should be pointing to this position.
- "FILL WATER" As long as the knob is in this position, the inner chamber will slowly fill with water.
- "STE" This position stands for sterilizing, and in this position, the autoclave will clean the samples.
- "EXH+DRY" In this position, the autoclave will do two things. The first is that it will neutralize the inside pressure with the outside pressure and then it will dry the samples.
- The three lights are:
- Power (top green) This light will be on when the autoclave is on.
- Dry (middle orange) This light will come on when the autoclave is drying the samples.
- Heat (bottom orange) When the autoclave is sterilizing the glassware, this light will come on.
How to use the Autoclave
Before using the autoclave, make sure that there is enough water in the reservoir. To do this, pull out the dipstick on the right side of the autoclave and look on the inside half of the strip. The indicator will be wet, and there should be enough water to wet the stick somewhere between the "Min." and "Max." positions. If it is low, fill it up with some DI water, but be careful to not exceed the maximum volume.
Sterilizing the Samples
- Mark everything that you are putting into the autoclave with the heat sensitive tape. The tape should be in the drawer under the autoclave.
- Place everything on the tray with some room between each item. If you are sterilizing liquids, use the non-perforated pan.
- Flip the green switch, upper right corner, to the "START" position.
- Turn the bottom knob, clockwise, to the "FILL WATER" position.
- Watch the water fill in on the inside of the cavity, when it reaches the front of the cavity, the small notch, turn the bottom knob to the "STE" position.
- Close the door and tighten it.
- Set the temperature knob, second from top, to your desired temperature.
- 273°F for plastics and other solids.
- 212°F for the liquids.
- Set the time, top knob, to the desired time.
- For plastics and solids 45 mins.
- For liquids 60 mins.
- NOTE: If the autoclave is already warmed up, subtract 10 mins from these times.
- The orange heat light, the lowest light, will turn on and the autoclave will run on its own.
- After the autoclave has ran through its cycle it will ding to inform you it is done. Depending on what you want to do follow one of the steps below.
Drying the Samples
- Wait for the pressure sensor to register 0 psi pressure difference between then inside of the autoclave and the lab. This is indicated by the white pointer on the upper dial pointing to 0.
- Slowly loosen the door about an inch to release the steam. Leave the door open to allow the steam to be released throughout the drying cycle.
- Turn the bottom knob to "EXH + DRY"
- Set the timer to ~20-30 mins.
- To indicate that the autoclave is working, the orange dry light, the middle light, will come on.
- After the cycle finishes, turn the bottom knob to the "0" position, open the door and remove samples.
- Flip the green switch to stop when you are done.
Not Drying the Samples
- Immediately turn the bottom knob to the "EXH + DRY" position.
- Wait until the pressure drops to 0 psi.
- Turn the bottom knob to the "0" position.
- Open the door, and remove the samples.
- Flip the green switch to stop when you are done.
Liquids
- Leave the bottom knob on the "STE" position, and wait for the pressure to drop to down to 0 psi.
- Turn the bottom knob, clockwise, to the "0" position.
- Open the door and remove the samples from the autoclave.
- Flip the green switch to stop when you are done.
REMEMBER: The autoclave is hot, very hot, so anything you put into it is also going to be hot when you are done with it. That being said, always use caution taking things out of the autoclave, you don't want to get burned, drop something or spill any liquids on you lap.
Cleaning the Autoclave
If you ever need to clean the autoclave, refer to this wiki entry for the procedure on how to clean it. It is recommended by the manufacturer to clean the autoclave once a week, or every twenty uses, which ever one comes first.
