There are parameters which are important to control the outcome of sonication, namely size of the probe, temperature, watt power used, and length and periods of time used during sonication. If you want to only crack the nuclear membranes in order to release the interacting proteins for immunoprecipitation study, then you should use a small probe, at 4 degree C, medium watt power, for 10 min X 3.

Thank you.

I would avoid sonication if possible as I believe it often damages proteins. A published study notes that sonication can result in formation of aggregates similar to amyloid (Protein Sci. 2004 November; 13(11): 3017–3027). The temperature at the probe is high and you are putting a lot of energy into your sample.

One way you can avoid the denaturation of proteins during sonication is what I mentioned above. The icy beaker with salt in it to lower the temp and maintain the temp at 4 degree C constantly, is able to absorbe the extremely high energy generated during sonication, while using a medium probe size and sonicate only briefly 10 min and repeating 3 times with 10 min cooling intervals to maintain at 4 degree C. Oh, this process should be in the presence of 1% triton X-100 to solubilize the cellular membranes.

Yes - cooling certainly reduces the damage. However, cooling the bulk liquid does not change the fact that ultrasound unavoidably produces a heat gradient from the probe. Protein diffusion and interactions are on the nanosecond scale so a 10 minute sonication allows plenty of time for sample damage.

For nuclear protein IPs I slightly modify the following 1%Triton X buffer given below and lyse the cells in 500µl IP-lysis buffer. Add 25µl of 5M NaCl, mix gently by flipping the tube. (20 min, on ice).

Add 500µl ice-cold water to dilute the salt. Preclear @ max speed (17,000 g) 20 min 4C and use the SUP for IP.

50 mM Tris HCl pH7.4

150 mM NaCl

1% TX

2mM NaF

1 mM NaVO3 (add freshly)

1x PI (protease inhibitor coctail roche, add freshly)