Why doesn't our autoclave sterilize our broth?
April 23, 2012 3:18 PM Subscribe
Why is weird stuff growing in my flasks of 1% tryptone broth after I've autoclaved them to mix and sterilize the broth?
posted by pwb503 to science & nature (14 answers total) 2 users marked this as a favorite
We routinely make 1% tryptone broth solutions in glass flasks, then to completely mix the broth and sterilize it, we put it in the autoclave. We set the temperature to 255 F and run it for 25 minutes before allowing it to cool/vent for 10 minutes. Since the liquid is boiling, to keep it from blowing up the flask, we unscrew the lid a bit so that it is still on but loose enough to allow pressure to vent from the flask. Additionally, we use "autoclave tape" to make sure the proper temp/pressure has been met. When the autoclave has run its course, we open it up, screw the caps on and call it good. There are further steps in the process but that's not germain to my current issue...
Within a few days (as few as 2 and as many as 7) something starts to grow (bacterial? yeast?) in what should be sterile broth. We've tried two different autoclaves. We have not yet tried new tryptone powder (but are about to...), nor have we tried a new source of DI water (but are too about to...) Additionally, we've tried upping the time in the autoclave to 35 minutes with no time for venting. The problem continues.
Does this sound familiar to anyone? Has our tryptone or water been contaminated with something that can survive the autoclave?