Differences between sonicators?

[Mindbomb]

Hi

I'm new here and looking for advice on fragmenting chromatin. We plan to do quite a bit of ChIPseq to identify transcription factor binding sites. We are not aiming at high-throughput work with plates etc. Instead, we work mostly with limited amount of cells (human blood or tissue samples). Hence our primary concern is to maintain the high quality of DNA during shearing and prevent sample loss. The aim is to go for Illumina sequencing.

I would be very glad if you could give some advice on sonicators. The one that we have at the moment is probably not suitable for the purpose (a large open 9l waterbath, 230V 50Hz 1100W). I have been looking around and all the suppliers can demonstrate nice gel pictures. I would really appreciate to hear real life user experience. The brands/models that I found are

*covaris
*EpiSonic Multi-Functional Bioprocessor 1000
*EpiShear Multi-Sample Sonicator with chiller
*Epishear probe sonicator with cooling device
*bioruptor

We will at most do 10 samples at a time, so I wonder whether some of the devices would be an over-kill for us? The prices are really different.

Is there a big difference in water bath based devices (ie the bioprocessor) versus cup horn like devices?

Thanks a lot for help!

[mudshark]

We have used the Bioruptor for years. However, since we switched from ChIPchip to ChIPSeq we got increasingly unhappy with it. I guess it is due to increased shearing as we needed smaller fragments for library preparation (700 bp for ChIPchip, 200 for ChIPSeq). So we had to shear using 50-60 cycles instead of 15-25. Apparently the/our bioruptors did suffer and had increasing performance problems after some time of usage (i.e. after few months the initial good performance was completely gone). Unfortunately the company does not really have a clue what is/was going on. They were very helpful replacing instruments but we were not happy with the overall unstable situation. If you consider the bioruptor I would strongly recommend to talk to bioruptor users that have roughly similar protocol and sample turnover that you will have. If you can foresee that machine running time will be little, you might consider to obtain it, if you plan 24/7 usage, I would recommend to look for alternatives.

We recently got a Covaris S220. Chromatin prepared with it can be very very nice. Best 200-250 fragments ever seen. However, at present the reproducibility is far from what company promised. But we are trying to find the source of variation. In addition, the S200 machines are single sample processors. We have 30 mins of shearing, i.e. with more than 5 samples chromatin fragmentation gets a bit annoying. Multiple sample Covaris machines are far too expensive for us.

[cement_head]

Hello,

I am curious as to why so many people are using Covaris as opposed to a handheld rig like this: http://shop.midsci.com/productdetail...des_6mm_probe/

Is this because of reproducibility? Protocols? The price difference is about $20,000; which is not trivial.

I am completely new to sonication for NGS protocols, so any advice would be helpful.

Thanks,
Andor

[kwaraska]

The most obvious difference is the ability to walk away. A Covaris-you put the tube in-hit start and walk away. For a sonication of 30 minutes-that alone is huge.

In addition, any time you introduce humans into the equation, the results will be less reproducible. A hand held sonicator is going to be different in every persons hands-literally and figuratively.

We have customers that used to use a probe sonicator and is completely switched over to Covaris after seeing the results.

As for the comment about reproducability, you should look at the Covaris website. We just hosted a seminar detailing this exact topic. The outcome is that the same cell line cultured differently will shear differently, so once you determine the best conditions (usually a time course) you can stay with those as long as you don't change the growth conditions.

[cement_head]

Ok, thanks - that's what I suspected. It would really be a huge disappointment to destroy samples due to bad sonication, and/or inability to replicate sonication.

[NextGenSeq]

Probe based sonicators give inconsistent results and heat the sample. The Covaris gives far more consistent results and doesn't boil your sample.

[qsonica]

new shearing device options

Qsonica now offers a high through-put Dna shearing system. Works as well as the competition and costs less. Sorry for advertising here but its a new option to consider. Can't hurt to check it out and speak to some references.

[bilyl]

Hello,

I am curious as to why so many people are using Covaris as opposed to a handheld rig like this: http://shop.midsci.com/productdetail...des_6mm_probe/

Is this because of reproducibility? Protocols? The price difference is about $20,000; which is not trivial.

I am completely new to sonication for NGS protocols, so any advice would be helpful.

Thanks,
Andor

If you're doing anything more than one sample, this gadget will be a big pain.

[cement_head]

If you're doing anything more than one sample, this gadget will be a big pain.

Yeah, that's the conclusion I came to after speaker with a number of genome centres - the reproducibility from sample to sample is a major concern.

- CH

[maxell]

but still I cant see any comments about differences between cup horn and water bath ? . which one is better do you think ? and is it comparable with the multi tip horns or cup horn is better than that ? thanks

[qsonica]

Water baths send random sound waves through the tank and generally give inconsistant results. Standard water baths operate at 40kHz which is low intensity and too weak for sample processing. These are really made for cleaning.
Cup horns are made to deliver sonication directly to the area where the sample tubes are located plus cup horns offer adjustable intensity settings. You can dial in the setting needed for your particular sample.

Newer high intensity cup horns are now sold specifically for ChIP and DNA shearing and work very well.

[maxell]

so can we say that multi probe like 96 tips are not effective as cup horn ?

[qsonica]

It is application specific. For ChIP/dna shearing the answer is yes, cup horn is much more effective.

[maxell]

do you have any idea how we can test q800r in europe ?

[crocky]

I have been using the Covaris (S series) for my chromatin fragmentation and I am very happy with it. It works very fast and reproducible. For the fragmentation I need 3 min, whereas I used to sonicate for more than 30 min with the Bioruptor. Therefore one can do multiple samples, even if you can sonicate only 1 sample at a time.