Results from collision energy optimization hober  2019-03-07

I am currently working with a Thermo TSQ Atis and have used the tutorial for collision energy optimization and this is truly an excellent tutorial.
I am as of now in the process of setting up a pipeline for screening a few hundreds of peptide standards we have in the lab and has run into some issues with the output from Skyline regarding the transition lists.

I have recalibrated the CE predictor based on my previous results and this works really well. However, when I run samples with longer peptides Skyline predicts collision energies up to 80, while the instrument only allows for collision energies up to 65. IS there any way of setting a global maximum value for Skyline?

An additional problem that I have run into is that Skyline occasionally chooses 0 as the collision energy for some peptides Even though I get good results for them in the CE optimization. Is it possible to tell Skyline to use the default value for the collision energy rather than 0 in these cases?

Another, more rare, issue that I've had is that Skyline sometimes provides me with duplicated Precursor-Product ion pairs which make Xcalibur complain. Is there any way around this?

I know that these are quite a few questions, but if you have any solution to any of the problems it would be much appreciated.

Best regards

Nick Shulman responded:  2019-03-07
I do not know the answer to your first question, but the others I think I could figure out if I saw your Skyline document.

Can you send us your Skyline document?

In Skyline, you can use the menu item:
File > Share > (complete)
to create a .zip file containing your Skyline document and supporting files including extracted chromatograms.

If that .zip file is less than 50MB you can attach it to this support request. Otherwise, you can upload it here:

-- Nick
Brendan MacLean responded:  2019-03-07
I will answer the first question: no such ability to set a maximum predicted value exists yet.

Sounds like a great idea, though, and relatively simple to add to the "Edit Collision Energy Equation" form. We could add "Min value" and "Max value" fields to the form. I guess we already enforce a min value of zero (exclusive). You are the first to request the maximum value enforcement.

Thanks for posting to the Skyline support board.

hober responded:  2019-07-16
Hi Nick and Brendan,
I know that this is a very late reply, but I completely forgot to send you the file.
I have now sorted out two cases of the issues I described above.

The first peptide in the document is a quite long peptide and one of the cases in which Skyline predicts a CE-value above the limit of the TSQ Altis (which is 65) and the second peptide is a case where two of the product ions end up having the same m/z.

I know that these problems might seem small, but unfortunately, this results in the method exporter not being able to export any method from the documents which include peptides giving rise to these errors.

Additionally, I want to thank you a lot for solving the issue with the export of methods for the TSQ Altis.