4 replies to this topic

[TracyDuke]

Hi All, I used to be able to have cells well sonicated in 1 ml solution. I am currently trying to do ChIP with less amount of cells hence in less volume solution (300 ul). I found the lysates get really dirty after sonication with dark precipitates. Why is that?

[cellcounter]

TracyDuke, on Jul 16 2009, 03:04 PM, said:

Hi All, I used to be able to have cells well sonicated in 1 ml solution. I am currently trying to do ChIP with less amount of cells hence in less volume solution (300 ul). I found the lysates get really dirty after sonication with dark precipitates. Why is that?

More information on cell type and machine you are using may be helpful.

[Radish]

TracyDuke, on Jul 16 2009, 02:04 PM, said:

Hi All, I used to be able to have cells well sonicated in 1 ml solution. I am currently trying to do ChIP with less amount of cells hence in less volume solution (300 ul). I found the lysates get really dirty after sonication with dark precipitates. Why is that?

Lately I am having the same problem  

In my case I am using a bioruptor 200, E14 cell line and they are being sonicated in 1 ml.

When I centrifuge the lysates after the sonication it looks like a thread, with brownish/black color.

Thank you very much in advance.

[KPDE]

Radish, on Nov 21 2009, 05:54 PM, said:

TracyDuke, on Jul 16 2009, 02:04 PM, said:

Hi All, I used to be able to have cells well sonicated in 1 ml solution. I am currently trying to do ChIP with less amount of cells hence in less volume solution (300 ul). I found the lysates get really dirty after sonication with dark precipitates. Why is that?

Lately I am having the same problem  

In my case I am using a bioruptor 200, E14 cell line and they are being sonicated in 1 ml.

When I centrifuge the lysates after the sonication it looks like a thread, with brownish/black color.

Thank you very much in advance.

Interesting.  I used to use a Misonix 3000 with a microtip and got what you're describing and then switched to using a Bioruptor and the dark precipitate went away.  In any case, I haven't seen much of a difference in yield or enrichment between when I was getting the precipitate and when it went away.

[Radish]

KPDE, on Nov 22 2009, 10:08 AM, said:

Radish, on Nov 21 2009, 05:54 PM, said:

TracyDuke, on Jul 16 2009, 02:04 PM, said:

Hi All, I used to be able to have cells well sonicated in 1 ml solution. I am currently trying to do ChIP with less amount of cells hence in less volume solution (300 ul). I found the lysates get really dirty after sonication with dark precipitates. Why is that?

Lately I am having the same problem  

In my case I am using a bioruptor 200, E14 cell line and they are being sonicated in 1 ml.

When I centrifuge the lysates after the sonication it looks like a thread, with brownish/black color.

Thank you very much in advance.

Interesting.  I used to use a Misonix 3000 with a microtip and got what you're describing and then switched to using a Bioruptor and the dark precipitate went away.  In any case, I haven't seen much of a difference in yield or enrichment between when I was getting the precipitate and when it went away.

I have been furiously trying to get into the fast-ChIP protocol and I can't even get passed the sonication.
Everything always goes completely wrong.
LOL the only way to alleviate my failure is to laugh    


Thank you so much for the feedback, at least this way I Know the it is not an unseen thing.

Edited by Radish, 22 November 2009 - 04:27 PM.