Techniques on sonication - Can anyone give me some suggestions? (Aug/23/2007 )

I have overexpressed a protein and prepared to purify it. I decided to use sonication to disrupte the cells. But I know that sonication can cause protein aggregation and can genetrate heat. How to optimize the yield of soluble protein by using sonication? Thanks

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sonicate for 2 sec and then put the tube on ice for 20sec.
repeat 5 times.
Avoid bubbles.

Hope this help.

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If you are lucky enough to have a variable power sonicator then this will also allow you to optimise the conditions. We have a MSE Sonicator with power output measured as amplitude microns. I think the scale goes from 0-20 Amp Microns and for our enzyme prep use half power (10 Amp Microns).
As minnie mouse suggested keep your sample on ice and sonicate for short bursts i.e. 5 second burst....25 seconds rest, 5 second burst......25 second rest. This allows the probe and samples to cool thus preventing denaturation of the protein.

Hope this is useful

Rhombus

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But is the sonication time volume dependent? I have 15ml of sample to deal with, so should the period of sonication be longer?

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I dont think it is a very good idea to sonicate the cells at a longer period. In general, longer time means more heat. Even shock burst technique might not helping at all.

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You can always split into 3 ml tubes. :-)

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I agree with genehunter.

I don't think you can efficiently sonicate the cells in 15 ml.
I think smaller volume is better.

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