Method for lysing cells for fractionation - (May/13/2009 )

Hello. I'm planning to do fractionation to determine the localization of peroxiredoxin. My organism is single-celled and I'm would like to avoid the use of detergents. We have a Dounce homogenizer in the lab, but I think percent lysis with this method will be low. Some people suggested to do freeze-thaw or to pass my cells through a syringe. Any comments or suggestion? Thanks.

If you have access to liquid nitrogen, a few freeze-thaw cycles (preferably in lysis buffer) should work fine. Dry ice may work too.

Most effective would be using a french press if you have access to one...

gfischer on May 13 2009, 07:00 AM said:

Thanks for the suggestion. We have liquid nitrogen in the lab, but won't freeze-thaw distrupt also the membranes of the organelles, eg. mitochondria?

mastermi on May 13 2009, 12:14 PM said:

Thanks for the suggestion but we don't have a french press in the lab. Is it OK to pass my cells in a syringe instead? I just want to distrupt the plasma membrane and not organellar membranes. Thanks again.