Sonicating vs. homgenizing - (Oct/05/2005 )

For tissue processing for WBs what is the difference of the resultant protein sample based on whether I sonicate or homogenize?

Thanks!

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USE SONICATION IN 0.1 M NaCl FOR 15 MIN

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I may be talking rubbish here, but I THINK that sonicating breaks doen the structure of the cell, allowing the cytosolic proteins out, and fragmenting the membrane, making it easier to get membrane bound protein out of the cell. (well that is the reason I sonicate!)

Homoginising is just mincing the tissue up really small isn't it? I remember doing it to rat brains in a blender because we wanted the cells to remain whole so we could look at the way the cell surface receptors reacted to radiolabelled LSD and MDMA. I still wonder how the got permission for undergrads to play with dangerous stuff like that!

Of course I might be wrong!

Rosie

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Hi, Rosie!
Do you lyse cells in boiling laemmli buffer before sonication? Or do you know a buffer that allows both to detect membrane-bound proteins and to quantify the total amount of proteins?
Thank you!
Nico

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Hi,

I think one added advantage of sonication is that it breaks down the DNA of the cells. This will reduce the viscosity of the sample for easier loading into the gel and will also enable better transfer of the proteins after SDS-PAGE.

Rgds,
Angela

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I don't lyse cells like that. I sonicate in an extraction buffer containing 10% Protease inhibitor cocktail the extract my cytosolic and membrane bound proteins seperatly using an ultracentrifuge. To seperate the protein from the membrane I use a buffer containing triton x100 as well as the pic.
Hope this helps.

Rosie

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