Importing negative mode MRM scan on Skyline (small molecule)

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Importing negative mode MRM scan on Skyline (small molecule) jung165  2019-02-28
 

Hello Skyline team.

I have a one quick question about importing negative mode scan of MRM in Skyline software (small molecule)
I am performing metabolite analysis with Agilent 6490 MS devices and export the collision energy optimization method on Skyline.

After analysis and when I import my raw files on Skyline, it gave me the error message shown below:
[Component Microsoft .NET Framework 4.7.2 (x86 and x64) has failed to install with the following error message:
"A failure occurred attempting to install the Microsoft .NET Framework 4.7.2."

The following components failed to install:

  • Microsoft .NET Framework 4.7.2 (x86 and x64)

See the setup log file located at 'C:\Users\Allen\AppData\Local\Temp\VSD86B0.tmp\install.log' for more information.]

I also used Skyline-daily version and latest version of Skyline but, all of results shows same error.
Is there any solution for this error?? please help...
I also attached my Skyline and raw files for the analysis.

I really appreciate for your help and look forward for your answer.

Sincerely,

 
 
Brian Pratt responded:  2019-02-28

Hi,

Have you tried installing the Microsoft .NET Framework 4.7.2 manually?
https://dotnet.microsoft.com/download/dotnet-framework-runtime/net472

I went ahead and tried working with the files you provided (thanks, very helpful!) and while I don't get that .NET error, I do get this:

At 8:21 AM:
Failed importing results file 'D:\downloads\Raw file\20190229_Mitocondria_TCA_Gly_negative_mode_02.d'.
This document contains only negative ion mode transitions, and the imported file contains only positive ion mode data so nothing can be loaded.

So while you have your document correctly configured for negative ion mode (that is, using [M-H]), it appears that the mass spec file doesn't actually contain negative ion mode data. When I view the data in SeeMS all the Q1 Q3 pairs are positive values.

You'll need to sort out the .NET issue first, but I'd also ask you to open that data in Agilent's tools to see if it's really negative ion mode or not, and we can go from there.

Best,

Brian Pratt

 
jung165 responded:  2019-02-28

Thank you Brian! Your response releases part of my curiosity

But, I am still trying to understand Q1 Q3 values are still positive.
I am kind new to negative mode scan and I thought only changing positive polarity to negative on Agilent acquisition list will gave me negative scan.
Do you have any advice for negative mode scan on Agilent 6490devices??

You mean that precursor/product ion m/z should be “negative value” on Skyline?? (Ex, precursor ion(m/z)= 171 -> -171 something like this)??

 
Brian Pratt responded:  2019-02-28

Do you have any advice for negative mode scan on Agilent 6490devices??

I don't have any idea about that, sorry.

You mean that precursor/product ion m/z should be “negative value” on Skyline??

No, your skyline file is fine for use with negative ion mode data since the adducts in use are all negatively ionizing. The problem is that the .d file you're trying to import isn't negative ion mode data, so Skyline finds no matches between its targets and the mass spec data. I used the ProteoWizard SeeMS viewer to inspect the .d file, it indicates that the data was not acquired in negative mode.

It's possible, of course, that SeeMS and Skyline are both confused about the .d file, but it would be best to inspect it with Agilent's own tools to convince yourself that it's really negative ion mode data.