best method for tissue disrupting

Dr Engelbert Buxbaum engelbert_buxbaum at hotmail.com
Fri Dec 20 09:40:26 EST 2002


Huseyin Kucuktas wrote:

> 
> What is the best method for disrupting tissues (muscle, liver) for
> subsequent enzyme analysis in native gels? Mechanical disruption?
> Sonication? Freeze-thaw? Any idea?

For your application you can probably use any of these methods, as you
are working with small amounts and do not require the maintainance of
subcellular structures. A small Potter or a sonicator are probably
easiest. 

Just remember to keep everything on ice during homogenisation, as both
methods produce heat.

Add a mix of protease inhibitors (1 mM PMSF (fresh from 1000 times stock
in DMSO, toxic), benzamidine, EDTA and e-aminocaproic acid (100 uM
each), pepstatin and leupeptin (2 uM each)).



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