Suitable lysis buffer to extract transmembrane protein? (eg. tyrosinase) - (Jan/16/2008 )
QUOTE (sasoriza @ Jan 17 2008, 07:42 PM)
Which type of protease inhibitor cocktail should be used for protein extraction from animal cell line? Use the one for general use (lyophilized powder form) or the one specific for mamamlian cell lysis (DMSO solution form)?
You could use any of them. I would prefer to use these than PMSF. Its difficult to dissolve PMSF (you have to dissolve in ethanol) and it works upto 30 min. and when you add it to the buffers, it precipitates. I am not really confident about it.
-scolix-
QUOTE (scolix @ Jan 18 2008, 06:20 AM)
QUOTE (sasoriza @ Jan 17 2008, 07:42 PM)
Which type of protease inhibitor cocktail should be used for protein extraction from animal cell line? Use the one for general use (lyophilized powder form) or the one specific for mamamlian cell lysis (DMSO solution form)?
You could use any of them. I would prefer to use these than PMSF. Its difficult to dissolve PMSF (you have to dissolve in ethanol) and it works upto 30 min. and when you add it to the buffers, it precipitates. I am not really confident about it.
The protease inhibitor cocktail (lyophilized powder form from sigma) for general use contains AEBSF, E-64, bestatin, leupeptin, aprotinin and sodium EDTA. It is water soluble and i think it will be easier for me to prepare the lysis buffer containing 0.1 % triton-X.
How to prepare the lysis buffer? Prepare the stock solution 1st? Then only prepare the concentration needed and mix into volume needed from the stock solution? All the solutions can stored at -4 C for how long?
-sasoriza-