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Method for lysing cells for fractionation


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#1 Tenuissimus

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Posted 13 May 2009 - 06:55 AM

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.

#2 gfischer

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Posted 13 May 2009 - 07:00 AM

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

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Posted 13 May 2009 - 12:14 PM

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

#4 Tenuissimus

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Posted 13 May 2009 - 07:10 PM

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


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?

#5 Tenuissimus

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Posted 13 May 2009 - 07:13 PM

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


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.




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