How to use Skyline to optimize MRM parameters

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How to use Skyline to optimize MRM parameters z15839328872  2024-10-08 01:48
 

Dear Skyline Team,
I'm having some issues with Skyline to optimize MRM parameters, and I don't know how to automatically optimize collision energy and declustering voltage. Since my previous protein and peptide identification was done by sending samples, using Thermo Fisher Fisher Scientific's fluid, the raw data was imported into Skyline and the ion pairing information was exported. Later, I used my own Agilent instrument to verify that the peak was too low, and I wanted to optimize the MRM parameters, but I haven't found how to use Skyline optimization, do you have any good suggestions? Or am I using a 2023 version that needs to be updated?
Zhou Yanan

 
 
Nick Shulman responded:  2024-10-08 02:24
Here is the collision energy optimization tutorial:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_optimize_ce
The collision energy optimization tutorial is only available in English.

The latest released version of Skyline is 24.1. We always recommend using the latest version of Skyline unless you are trying to reproduce results from older experiments.
I can see from your screenshot that you are using an older version of Skyline because Skyline 24.1 uses different images for the green, yellow and red peak quality dots that you see next to the items in the Targets tree.
Nothing has changed in a long while in the area of SRM collision energy optimization, so the collision energy optimization tutorial will work with older versions of Skyline. However, we do recommend that you upgrade to Skyline version 24.1.

By the way, I see that many of the transitions in your Targets tree have no green, yellow, or red symbol next to them which usually indicates that there was no matching chromatogram in the data file. This often happens if your data was collected across multiple injections.
When you do "File > Import > Results" in Skyline, the first window that comes up has some options which would allow you to combine multiple injections into a single Replicate. I have circled those options in the attached screenshot.

You might also find some useful information in the Targeted Method Refinement tutorial:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_method_refine
this tutorial is also available in Chinese:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_method_refine_zh

-- Nick
 
z15839328872 responded:  2024-10-08 05:27
Thank you for your reply