Requests

support
Showing: limited to 100 requests
Bug in document grid reporting
(4 responses) jmeyer 2020-07-08

Dear Skyline team,

I believe I found a bug in skyline reporting identifications in runs where the peptide was in fact not identified.

Please see the two attached screenshots where there is clearly no identification line in the chromatogram, but the document grid reports "TRUE" for an identification in that run (top line in the document grid). These are two separate peptides. When I manually drag the chromatogram to re-integrate anywhere in that run, even the same area, it goes to FALSE.

Example 1 is R.SEQEDEVLLVSSSR.Y [27, 40] ++

Example 2 is R.YPDQWIVPGGGMEPEEEPGGAAVR.E [41, 64] +++

I also attached a minimal skyline share with these two peptides, and the other peptides in the same protein.

Any help fixing this or providing a workaround would be greatly appreciated.

Best regards,
Jesse

 Inkedwrong ID 1 skyline_LI.jpg  wrong ID 2 skyline.JPG  wrong_IDs_example_small.sky.zip 
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Quantifying on multiple fragment ion isotopic peaks
johan gobom 2020-07-08

Dear Skyline Team,

I’m setting up a PRM method for large peptides, around 4.5 kDa, +4-charged, on an Orbitrap. For many of the fragments, the monoisotopic peak is not the strongest by far, so I would like to quantify on up to four isotopic peaks for each fragment. Is this possible to do in Skyline?

All the best,

Johan

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Making libraries for synthetic peptides and packaging them into single file in skyline
(2 responses) SRS 2020-07-07

Hi There,

I have been trying to make a spectra library of about few thousands synthetic peptides from a DDA run. I was able to import the data using "mziden" and "mzxml" files from the search and mass spectrometry data, respectively. But I have two questions here, 1) how can I export all the spectra libraries into a single file in skyline?

  1. How could I do the peak picking for over thousand peptides, as they are so many, it is not possible for me to open them to make sure the RT is aligned.

Do I need to make sure the alignment of the RT is correct then package them?

I can send you the skyline file if that help,

Thank you, looking forward to hearing from you.

Regards,
SRS

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Failed to export method for CE optimization
(10 responses) benoit fatou 2020-06-08

Dear Skyline Team,
I tried to export a MRM method to perform CE optimization and I have got this error message (see attached file).
Could you please help me solve this issue?
Thanks,
Best,
Benoit

 ErrorSkyline.PNG 
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TMT0 Labeled Peptide as Internal Standard for TMT10 Targets
drothenberg 2020-07-07

Instead of using a heavy labeled synthetic peptide as an internal standard, I'm interested in using a TMTzero labeled peptide to compare against TMT-10plex labeled targets. I know ratios can be calculated using the Surrogate Standard option, but is it possible to have synchronized zooming and transition matching similar to heavy-labeled peptides? This would be similar to the TOMAHAQ approach, but without MS3. Thanks.

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QuaSAR Error
msj0506 2020-07-06

Dear Skyline Team,

I have a problem using QuaSAR tool in Skyline.

Now, my desktop system is window 10 and QuaSAR tool is connected R 4.0.0 ver in Skyline.

But, When i used QuaSAR for calculating LOD and LOQ, the error occurred continuously. i attach the related file.

Reinstallation does not solve the problem, and the same problem occurs in my laptop. Can you tell me how to solve this?

 Error(1).PNG  Error(2).PNG 
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Calibration curve calculations problems.
(2 responses) monika lysakowska 2020-07-06

I have a problem calculating the calibration curve. The calculations Skyline does are different from my calculations in Excel.​I have entered all the appropriate values for settings and grids
and I got a good calibration curve. R square and intercept is identical to the calculations in the excel spreadsheet, slope differs by two decimal places (I think it is related to the Y-axis conversion). However, the calculated values by Skyline Quan differ slightly from heavy normalized to light values from the excel worksheet.

The data should match the normalized skyline. Currently they are about a percentage point off, which is substantial. It’s about 20% difference between the two. What can explain this?​

Regards,
Monika Lysakowska

 1.png  2.png  3.png 
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Install issue
(3 responses) Brett Phinney 2018-01-30
Hey everyone, not sure if this is just my messed up win10 computer but I tried to install

https://skyline.ms/software/Skyline-release-64_4_1/setup.exe

And win10 would not let me install it even as the admin.

This seemed to fix it

https://superuser.com/questions/1252575/unable-to-install-clickonce-application-due-to-security-settings-windows-10


just an FYI if someone else has this issue

Cheers

Brett
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White screen on Collision Energy Regression
(1 response) benoit fatou 2020-07-03

Hi Skyline Team,

I am doing some CE optimization on a Shimadzu instrument.
After editing the Collision Energy Equation, I would like to visualize the corresponding graphs but I have a white screen after clicking on "Show Graph...".
I have the same issue when I select default equations from other instrument vendors.

Let me know if you need more information and I am happy to send you the Skyline document.
Thanks,
Stay safe!
Benoit

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Failure opening
(1 response) Flora.Sh. 2020-07-03

Dear Skyline Team

At my workplace I work a lot with skyline. If I want to work at home at my laptop there is a problem. I can not open the files because it says that my document format version is 20.13 and newer than the version 20.1 supported by skyline (64bit) 20.1.0.155 (a0e7323e3), which I have in my laptop. How can I change that or what can I do? There is not a newer version at your homepage.

Thanks in advance for the answer

Best wishes

F.Sh.

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Lipid isomer data import problem in scheduled MRM
(4 responses) stolltho 2020-07-02

Hi sky team.

A similar problem was reported earlier (Small Molecule MRM extraction with isomers) but it seems the problem persists.

I've got a long list of lipids, some of which have the same transition pairs (774.6>184.1).
Following scheduled MRM data import, skyline is showing only one / the same sMRM window for all the lipid isomers.

I already changed product ion m/z by 0.01 (774.6>184.10, 774.6>184.11) to circumvent the problem and it worked, but unfortunately the QQQ we are using is not acquiring identical peak profiles for this slight shift.

Hence I changed product ion m/z by 0.0010 (774.6>184.1000, 774.6>184.1010, 774.6>184.1009 etc.), but now I face the same import issue again, showing only one / the same sMRM window. I changed match tolerance to 0.0001, but then no chromatograms are shown after import.

Cheers,
Thomas

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different heavy IS concentration
(3 responses) Wael 2020-07-02

Dear Skyline team,
as we do multiplexed analysis we might need to adject the amount of spiked IS standard depending on the endogenous peptide level to be within a defined (SIS/EN) range. Is it possible to feed the results grid with different IS standard concentration for each peptide individually, or we should create a different skyline folder for each analyte to be able include the particular SIS concentration?

The issue also applies when creating a standard curve using range of different EN concentrations since the dilution curve concentration range vary depending in the analyte.

Best regards
Wael

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Import of Mascot error tolerant searches in Skyline
ingo.wohlgemuth 2020-07-02

Hey Skyline team,

I try to import error tolerant Mascot searches with a focus on amino acid substitutions. While many PTMs can be imported, amino acid substitutions which were found in Mascot are often lost in Skyline (e.g L to V mutations, see picture). Is the a rational for that? How Sykline handles cases is which the delta mass could not unambiguously assigned (e.g: D to E substitution vs D methylation) when both interpretations are available in Mascot?

Thanx a lot!
Ingo

 Amino acid substitutions.jpg 
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collision energy prediction for QTRAP 6500+
(1 response) mark adams 2020-07-01

Hey Brendan,
We are currently using a SCIEX QTRAP 6500+ platform for protein-based work. Prior to transition list export, would a collision energy prediction setting of 'SCIEX' or 'ABI 5500 Q Trap' be more appropriate for our instrument platform?
Thanks for any input!
-Mark

 Annotation 2020-07-01.png 
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SAV recognition by speclib builder
(1 response) dkueltz 2020-06-27

Hi Brendan et al.
I am using SPIDER within PEAKSsuiteX to identify allelic variants that manifest themselves at the protein level as single amino acid substitutions (SAVs). These can be exported from PEAKS to pep.xml + mzxml files that Skyline uses to built spectral libraries from DDA data. However, unlike unmodified peptides and regular PTM peptides, the SAV peptides are not recognized by the spec library builder. I was not able to open the mzxml files but the structure of SAVs is a bit different from that of regular PTMs in the pep.xml files (see below). I would be happy to send you a pair of pep.xml + mzxml files that can be used for speclib construction with Skyline and contain umnmod. PTM, and SAV peptides. Is there any way that I could import these SAV peptides into Skyline spectral libraries such that I can quantify the stoichiometry of allele-specific expression (ASE) in different biological contexts? If this is not possible yet, do you have any plans to implement this capacity into Skyline in the future?
Thanks,
Dietmar
Below is part of a pep.xml file that shows 3 entries, one each for a regular PTM, SAV, and unmod. peptide... in this particular example the PTM and SAV are actually the same - deamidation of Q to E - but there are many other instances with unique SAV mass changes that do not match a conventional PTM and can be interpreted unambiguously...
<spectrum_query spectrum="DK0020-B1B_2-A,1_01_2365.d" start_scan="2626" end_scan="2626" precursor_neutral_mass="1515.7789" assumed_charge="3" index="1303">
<search_result>
<search_hit hit_rank="1" peptide="LSNGTTKPQTNGVAK" calc_neutral_pep_mass="1515.7893" massdiff="4.7" num_tot_proteins="1" protein="XP_033982937.1" protein_descr="mucin-5AC [Trematomus bernacchii]" protein_mw="124905.71">
<modification_info modified_peptide="LSNGTTKPQ(+.98)TNGVAK">
<mod_aminoacid_mass position="9" mass="129.04259"/>
</modification_info>
<search_score name="-10lgP" value="44.82"/>
<search_score name="confidence" value="0.99"/>
<search_score name="PeaksScore" value="99.8"/>
<analysis_result analysis="reconstruction">
<tags denovo_tag="LSNGTTKPQTNGVAK" reconstructed_tag="LSNGTTKPQTNGVAK"/>
</analysis_result>
</search_hit>
</search_result>
</spectrum_query>
<spectrum_query spectrum="DK0020-B1B_2-A,1_01_2365.d" start_scan="2935" end_scan="2935" precursor_neutral_mass="1515.7789" assumed_charge="3" index="1304">
<search_result>
<search_hit hit_rank="1" peptide="LSNGTTKPQTNGVAK" calc_neutral_pep_mass="1515.7893" massdiff="4.7" num_tot_proteins="1" protein="XP_033982937.1" protein_descr="mucin-5AC [Trematomus bernacchii]" protein_mw="124905.71">
<modification_info modified_peptide="LSNGTTKPE(sub Q)TNGVAK">
<aminoacid_substitution position="9" orig_aa="Q"/>
</modification_info>
<search_score name="-10lgP" value="41.80"/>
<search_score name="confidence" value="0.95"/>
<search_score name="PeaksScore" value="95.8"/>
<analysis_result analysis="reconstruction">
<tags denovo_tag="LSNGTTKPETNGVAK" reconstructed_tag="LSNGTTKPETNGVAK"/>
</analysis_result>
</search_hit>
</search_result>
</spectrum_query>
<spectrum_query spectrum="DK0020-B1B_2-A,1_01_2365.d" start_scan="2596" end_scan="2596" precursor_neutral_mass="1088.5857" assumed_charge="3" index="1309">
<search_result>
<search_hit hit_rank="1" peptide="YTRPTPVQK" calc_neutral_pep_mass="1088.5978" massdiff="0.4" num_tot_proteins="1" protein="XP_034007142.1" protein_descr="ATP-dependent RNA helicase DDX3X-like [Trematomus bernacchii]" protein_mw="80330.54">
<search_score name="-10lgP" value="41.08"/>
<search_score name="confidence" value="0.97"/>
<search_score name="PeaksScore" value="96.3"/>
<analysis_result analysis="reconstruction">
<tags denovo_tag="YTRPTPVQK" reconstructed_tag="YTRPTPVQK"/>
</analysis_result>
</search_hit>
</search_result>
</spectrum_query>

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Issue: The following modifications could not be interpreted
(3 responses) Mark Athanason 2020-06-29

Hello Brendan,

I saw another post on your forums from someone with a similar issue, but mine is different. Long story short I performed a search using Comet with a few variable modifications on K,R,S,T,C (drug adducts) along with oxidation of M. When I import my data into skyline to build a spectral library (using the wizard) the Add Modifications window lets me pick one of my adducts and the oxidative M. These are rounded to the nearest one's place. The modifications that cannot be interpreted have 2, 4, and 5 decimal places. In my comet parameters file I have the mass going out to 4 or five decimal places for all variable mods. If you can let me know how to circumvent this using the search data rather than manually entering the variable mod in skyline (future up-scaling of the project), I would appreciate it.

-Mark

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Skyline document with cross-linked peptides
(3 responses) danielacgranato 2020-06-27

Sorry, I forgot to attach the documents in the prior message (pLINK output and prox.xml converted). Thank you very much. Best, Daniela

 plink_12_maio_2020.zip  proxl.xml_convert.zip 
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Different retention times for heavy and light from
(2 responses) Martin Eisinger 2020-06-23

Hi,

I know the topic has been discussed several time but I didn't find a conclusive answer. I work with small molecules and use fully deuterated versions as internal standards. The deuteration leads to a considerable RT shift between heavy and light form. However if I define them as heavy and light in my transition list, skyline uses the same integration boundaries for both forms. The only solutions I found to overcome this problem is to set very wide integration boundaries, so that both peaks heavy and light are covered. It's ok but no perfect solution as in that case a lot of baseline is also integrated.

Is there any good way of telling Skyline that heavy and light RTs don't have to match and allowing it to use different integration boundaries for both forms?

Thanks in advance!

Cheers,

Martin

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Quantifying labelling efficiency
(2 responses) sudip ghosh20975 2020-06-24

Hi I have a set of reference peptides which I labelled with TMT0. I ran both the labelled and unlabelled samples in Fusion and wanted to evaluate the labelling efficiency in Skyline. How do I proceed?

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SkylineRunner.exe to export mProphet Features
(6 responses) s26guan 2020-06-25

Dear Skyline team,

Can you show me the export option for mProphet features? I tried --report-name="mProphet Features" and it was not working.

Got error: "Error: The report mProphet Features does not exist. If it has spaces in its name, use "double quotes" around the entire list of command parameters."

Many thanks.

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Average peptide masses instead of monoiso
(1 response) Zac 2020-06-25

Is there a way to work with average peptide masses instead of monisotopic masses in skyline

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Reprocessing with new parameters
(2 responses) thomas boegl 2020-06-24

Hello.
I have following problem:
When I change any parameters e.g. adding new molecules ( Insert --> Transition List... ) I do not get any results until I do "Manage Results.." and Re-Import the files. Is there a simpler way to do reprocessing only for the new molecules?
I am currently starting to work with skyline.

Thank you!

Thomas

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Problem performing iterative CE optimization with explicit collision energies
(6 responses) vincentroyrichard 2020-06-10

Hi!

I'm currently trying to perform iterative CE optimization for an MRM method on a QTRAP 6500+ instrument using the latest version of skyline daily. What I would like to do would be 3 rounds of CE optimization with successively decreasing step sizes (starting with the CEs derived from the linear equation). What I'm noticing is that for the second pass CE optimization, I can only export a transition list for CE optimization with the optimized CEs (ie not derived from the linear equation) when entering these CE values in the explicit collision energy tab in the documents grid. The problem is that when I'm reimporting the data from the 2nd pass optimization it doesn't "center" around the correct CE.

For example, after optimization of a single MRM transition, the optimal CE was determined to be 18.6. I then reanalyzed the sample with a method to optimize the transitions with 3 steps +- 2 (ie 12.6 , 14.6, 16.6, 18.6, 20.6, 22.6, 24.6 -with 18.6 still remaining the most sensitive) - however when I reimport the data from this into skyline it changes the center CE to 24.6.

Seems to be a bug? Any assistance with this would be greatly appreciated!

Thanks very much!

Vincent Richard

 troubleshoot trans list.csv  troubleshooting.sky.zip 
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dotp / rdotp between samples
(2 responses) Juergen 2020-06-23

Dear Skyline team,

I currently use PRM in the identification of several peptides. My reference peptides are, however, not labeled and hence I measure them in separate runs. I know it is possible to get dotp values for each measurement in comparison to the corresponding library spectrum but I would prefer the dot product of the selected transitions in comparison to my control run (similar to the rdotp value for labeled and unlabeled peaks within one run). Is it possible to get this value?

I would like to use this value instead of the dotp against the library spectrum because I guess that than all the MS2 spectra below the peak would count for the calculation, while in the library only those spectra are retained that are nicely identified.

Best regards,
Juergen

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DIA method for Exploris 480
1kubo f 2020-06-23

Hi,
I would like to ask whether Skyline has a function to export a DIA method (MSX) for Exploris 480 operated by Xcalibur 4.3.73.11. Thank you, best regards Jakub

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Interesting error: TIMSTOF and Mascot
(4 responses) Jason Held 2020-06-12

Hi,

I'm trying to build a library from TIMSTOF data search with mascot. I'm using the DAT files to build the library and getting this error:

System.IO.IOException: ERROR: Maximum limit of 2000 spectrum source files was exceeded. There was most likely a problem reading the filenames.
ERROR:
ERROR: reading file F013408.dat
ERROR: Maximum limit of 2000 spectrum source files was exceeded. There was most likely a problem reading the filenames.
ERROR:
ERROR: reading file F013416.dat

Command-line: C:\Users\lab_held.pcl-held1\AppData\Local\Apps\2.0\NRM38HKN.2B5\OM37C28G.N7K\skyl..tion_e4141a2a22107248_0014.0001_1e739a267632795e\BlibBuild -s -A -H -o -c 0.99 -i Bowman005_TIMSTOF -S "C:\Users\lab_held.pcl-held1\AppData\Local\Temp\tmp2696.tmp" "C:\Skyline\SpectralLibraries\Bowman005_TIMSTOF.redundant.blib"
Working directory: S:\Users\Jason\Bowman005-New2CysMutantForPaperTIMSTOF
at pwiz.Common.SystemUtil.ProcessRunner.Run(ProcessStartInfo psi, String stdin, IProgressMonitor progress, IProgressStatus& status, TextWriter writer) in C:\proj\pwiz_x64\pwiz_tools\Shared\Common\SystemUtil\ProcessRunner.cs:line 62
at pwiz.BiblioSpec.BlibBuild.BuildLibrary(LibraryBuildAction libraryBuildAction, IProgressMonitor progressMonitor, IProgressStatus& status, String& commandArgs, String& messageLog, String[]& ambiguous) in C:\proj\pwiz_x64\pwiz_tools\Shared\BiblioSpec\BlibBuild.cs:line 201
at pwiz.Skyline.Model.Lib.BiblioSpecLiteBuilder.BuildLibrary(IProgressMonitor progress) in C:\proj\pwiz_x64\pwiz_tools\Skyline\Mode\Lib\BiblioSpecLiteBuilder.cs:line 155

Cheers,
Jason

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Issue uploading a Skyline Document to Panorama with a library file (.clib)
(2 responses) hogan 2020-06-18

Hello,

I have encountered an issue uploading a Skyline document to panorama that includes a .clib file. When I remove the .clib file using the peptide settings -> Library tab I am able to upload the file to panorama without issue. I would like to be able to include the .clib file in the document as the library dot product is data we would like to keep in the doc and it is not available if the library file is removed. Here is the error I receive from panorama when I attempt the upload. Thanks for your support!

18 Jun 2020 17:18:19,750 INFO : Starting to run task 'org.labkey.targetedms.pipeline.TargetedMSImportTask' at location 'webserver-high-priority'
18 Jun 2020 17:18:19,762 INFO : Starting to import Skyline document from 20200615_H9407_1280777_1280778_2020-06-18_17-14-33.sky.zip
18 Jun 2020 17:18:19,772 INFO : Expanding Popeye Chromatogram Library 20min_rev7.clib
18 Jun 2020 17:18:19,876 INFO : Expanding 20200615_H9407_1280777_1280778.skyd
18 Jun 2020 17:18:21,030 INFO : Expanding 20200615_H9407_1280777_1280778.sky.view
18 Jun 2020 17:18:21,059 INFO : Expanding 20200615_H9407_1280777_1280778.sky
18 Jun 2020 17:18:21,367 WARN : The version of this Skyline document is 20.1. This is newer than the highest supported version 4.1
18 Jun 2020 17:18:21,390 INFO : 1% Done
18 Jun 2020 17:18:24,287 INFO : Failed to complete task 'org.labkey.targetedms.pipeline.TargetedMSImportTask'
18 Jun 2020 17:18:24,314 ERROR: SqlExecutor.execute(); uncategorized SQLException for SQL []; SQL state [25P02]; error code [0]; ERROR: current transaction is aborted, commands ignored until end of transaction block; nested exception is org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block
Caused by: org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block
Caused by: org.postgresql.util.PSQLException: ERROR: insert or update on table "spectrumlibrary" violates foreign key constraint "fk_spectrumlibrary_librarysourceid"
Detail: Key (librarysourceid)=(0) is not present in table "librarysource".
18 Jun 2020 17:18:24,378 ERROR: Uncaught exception in PipelineJob: (DONE) Skyline document import - 20200615_H9407_1280777_1280778_2020-06-18_17-14-33.sky.zip
org.springframework.jdbc.UncategorizedSQLException: SqlExecutor.execute(); uncategorized SQLException for SQL []; SQL state [25P02]; error code [0]; ERROR: current transaction is aborted, commands ignored until end of transaction block; nested exception is org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block
Caused by: org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block
Caused by: org.postgresql.util.PSQLException: ERROR: insert or update on table "spectrumlibrary" violates foreign key constraint "fk_spectrumlibrary_librarysourceid"
Detail: Key (librarysourceid)=(0) is not present in table "librarysource".

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Case sensitivity for .mzID files
(2 responses) hayes.mcdonald 2020-06-19

When I am trying to import .mzID files from Morpheus searches, I end up having to change the suffix to all lower case mzid before Skyline will process. It isn't a huge deal, but it is annoying as the set of files gets larger.

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Metabolite ID Workflow for Oligonucleotides
(1 response) Richard Lam 2020-06-17

Can Skyline perform metabolite ID for oligonucleotides? If yes, any tutorials or webinars available. Thanks!

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Peak extraction window shifting
(4 responses) rmolden 2020-06-18

Hi,
My retention times shift very little from run to run, so I would like to use a stringent retention time filtering (0.5 minutes of MS/MS IDs) when chromatograms are extracted to reduce noise. When I import my results I notice that for some peptides the window in which XIC are extracted shift from run to run (see attached), even though the library is built just from one run and I have not applied any iRT calibration. Is there any way to make sure that that extraction window stays fixed across runs?
Thanks!
Rosie

 RT shift.PNG 
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Change the scale of the peptide intensity
(1 response) benoit fatou 2020-06-18

Hi Skyline Team,

I was wondering if it is possible to do a log2 transformation of the peptide intensities to then export the chromatogram as an image.
The idea is to uniform the intensity and make the chromatogram much nicer to visualize.

Thanks,
Best,
Benoit

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Retention time Prediction
(5 responses) scv01958 2020-04-23

Hi,

I had a question regarding the retention time prediction tool of Skyline. I understand that the tool is capable of using imported data to calibrate a calculator and build up an iRT database. This iRT database is then very useful in validating retention times recorded in future trials. However, I am confused on the "prediction" capabilities of this tool. Let's say I have a calibrated calculator already made consisting of an iRT database of related peptides/molecules. I have a new peptide/molecule that is related, but slightly different in some way, the difference is a change in structure or charge. Will this calculator be able to predict the retention time of my new compound based on the database I have already built?

Thanks.

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PRM - Unstable signals
(5 responses) melanieH 2020-06-15

Dear all,

I would like to have your advices regarding my issue.
We are working with a newly acquired Q-exactive HF-X and I am trying to setup a PRM method to quantify one bacterial protein (matrix : bacterial culture supernatants).
My issue is the signal that I obtain in scheduled PRM method, it seems really unstable but I am not sure if it comes from my method (actually pretty basic, following Thermo recommendations) which could be too much for just a low number of peptides [PRM R=30000, AGC t = 1e5, IT max = 60 ms, isolation window = 1.6m/z ) or if it could come from the instrument itself.
I attached a pdf file with some screenshots. I have more data but the results are similar for all the runs operated in scheduled mode, with or without FullScan. As you can see the signals seem "smoother" when the method is unscheduled.
I am asking if it could come from the instrument because we had once a problem with a lens in the quad that fell down during a cleaning session by the Thermo technician...

All advices are welcome.

Thanks

 2020-0615.pdf 
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Exporting resolution values
(4 responses) sstoychev 2020-06-16

Hi,

I would like to export the resolution values for both precursors and fragments in MRM-HR and DIA data but can't find the metrics in the report templates. There are FWHM and Max FHWM values but I think these are related to chromatography rather than mass spectra?

Regards,

Stoyan

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Probem in creating library from Maxquant results
(6 responses) nicolas pierre 2020-06-10

Hello,

I want to create a library from Maxquant results but I did not find the modifications.xml file.
So, I have an error message. Where to find the modifications.xml file ?
Thank you for your help,

Nicolas Pierre

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Issues with removing the display of compound names in skyline
(1 response) julien faugere 2020-06-16

How to remove compound names when overlaying multiple MRM transitions ?

 Capture.PNG 
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Prosit
(1 response) ingo.wohlgemuth 2020-06-16

Hey Skyline Team,

can I export individual prosit scores via the document grid?

Thanks a lot!

Best
Ingo

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Special SIL peptide standard library
(7 responses) lyjgbb 2020-06-10

Hi Skyline Team,

I wish Skyline could have a function to allow us to build a special SIL peptide standard library, which includes the customized stable-isotope labeled peptides as standards for our target LC-MS quantification on the unlabeled peptides from either cells or tumor samples. Because we are focusing on the HLA peptides, which are not typical tryptic peptides but they have a kind of diverse peptide sequences, we have a very large SIL peptide library with peptides labeled on diverse amino acids. When we run different samples, we have to choose different SIL peptides from the library, input the corresponding light partners first, and then manually modify the amino acids into SIL version one by one. So this process is very time-consuming and has risks for many human errors. It will be greatly helpful if we can set up a SIL peptide library first in Skyline and then choose the selected SIL peptides per unique sample efficiently from the library, by just inserting the light partners into Skyline. That means Skyline will give users a chance to choose which library (typical protein database or SIL peptide library) they would like to match the peptides with when they insert peptide sequences into the program.

Thank you so much for your great efforts and kind consideration!
John

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How to import amino acid substitutions from ProteinPilot 5.0 search results
(1 response) christof.lenz 2020-06-16

Dear all,

I am looking for a systematic way to import peptide search results from SCIEX ProteinPilot 5.0 including amino acid substitutions. In 'thorough' mode, ProteinPilot allows to search peptide sequences including all amino acid substitutions. Scoring is then adjusted using an underlying BLOSUM substitution matrix and goes into scoring, so we could use a peptide score cutoff here.

I am stumped, however, on how to import the search results including the 180 or so possible AA substitutions into Skyline to look for these peptides e.g. in HR-MS1 or DIA data. Is there any good way to do this, except 'declaring' each potential substitution as a variable PTM?
Data are from a SCIEX TT5600+ with Analyst TF 1.7; PP is version 5.0 rev. 4769; Skyline is on the latest version.

Any hints or experiences much appreciated!!

Chris

view request
Document Grid
Tobi 2020-06-15

Dear Skyline team,

hope you are doing well. I would like to drop some inspiration below.

Could you please add the Acquisition cycle time in seconds to the document grid? I know that people can calculate that themselves, but it would be very helpful to have the option for QC to check it directly in Skyline without having to export first and calculate by hand. The cycle time in seconds would be ((End Time - Start Time) / Points Across Peak * 60s) on the Transition level.

Another really important value was highlighted by the Heidelberg CF, which is the Tailing Factor. It is as old as chromatography itself and crucial in QC for observing tailing which requires hardware maintenance as well as indicating interference for reviewing transitions. It would work extremely well together with peak picking by avant-garde DIA as the boundaries could be set in an X% max intensity fashion. Please see below.

Page 27: https://skyline.ms/_webdav/home/software/Skyline/events/2020 User Group Meeting at ASMS/%40files/Presentations/1-01-Sakson.pdf

https://blog.sepscience.com/liquidchromatography/back-to-basics-5-tailing

With best regards,
tobi

view request
Is it possible to use the Skyline program for the quantitative analysis of various modified forms of the same peptide in different samples with known relative ionization coefficient k?
(1 response) akulyyasov 2020-06-12

Dear Brendan,
I have a question for you regarding the use of the Skyline program for the quantitative analysis of various modified forms of the same peptide in different samples.
According to the results of the experiment, I got two samples. In one control tube, a small amount of the biotinylated form of BAP is present, and basically there is a propionylated form of BAP.
In another tube, the amount of biotinylated form of BAP is much higher than propionylated BAP.
Does the Skyline program provide for the possibility of calculating the level of biotinylation between samples taking into account the known value of the relative ionization coefficient k of two modified forms of peptides?

With best regards
Arman


This Biotin Acceptor Peptide (BAP) is a non-naturally occurring peptide and is absent in the NCBI and Uniprot database.
BAP is the result of a design in which the amino acid sequence is modified to reduce substrate activity compared to the known natural AviTag peptide.

Biotin Acceptor Peptide (BAP) - tryptic peptide for LC-MS/MS
BAP biotinylated - ILEAQK(Bio)IVR, m/z value of (M+2H)2+ (648.4)
BAP propionylated - ILEAQK(Pro)IVR, m/z value of (M+2H)2+ (563.2)

The relative ionization coefficient k between the biotinylated and propionylated BAP peptides was estimated as k = (HP - LP)/(LB - HB), where HP corresponds to the area of TIC of heavy propionylated, LP of light propionylated, LB of light biotinylated, and HB of heavy biotinylated BAP. K were calculated for two independent experiments, giving an average value 11.9 (Kulyyassov A, et al. PUB-MS: a mass spectrometry-based method to monitor protein-protein proximity in vivo.//J Proteome Res. 2011 Oct 7;10(10):4416-27. doi: 10.1021/pr200189p).

I also uploaded one experiment in Panorama database

https://panoramaweb.org/Nat Center for Biotech Kazakhstan – Mass Spec Lab/BAPSx2_9hrbiotpulsonNidigest1_RA3_01_562/project-begin.view?

view request
Intact proteins+MRM
(2 responses) z1854947 2020-06-12

Hello,

I am new to the skyline family.

My current project is specifically focusing on Top down proteomics. I am separating and quantifying intact proteins using Shimadzu Triple-Quadrupole 8045 by optimizing MRM methods.

Can anyone help me by listing the specific tools/setting which will be helpful for my kind of work? Most of the tools on Skyline that I learnt are for tryptic digested peptide.

Also, on the instrument, my files are generated with .mzxlm extension. How can I upload these files?

Thank you.

view request
Full scan with MS1 filtering--- the intensity of precursor doesn't show up in the chromatogram
(4 responses) yusyuanluo 2020-06-08

Dear Skyline team,
I imported IM-QTOF data (Agilent) with a full scan setting as attached. I couldn't see the "peak" in intensity versus retention time plot.
However, the MS spectra do show up with intensity if I click on a time point in the intensity versus retention time plot. Is there any way that I can resolve this problem?

Thanks!

YS

 Skyline.pptx 
view request
Installation on Windows Server 2019
(5 responses) s26guan 2020-06-10

Dear Skyline Team,

Would you recommend a version to install on Windows Server 2019?

Thank you.

view request
Cross-link Transition Calculator - Problem/input
(4 responses) tatianibl 2020-06-10
Dear Skyline Team,
How are you?

I tried to use cross-link transition calculator tool and I am having a problem during the input process (error message below).

System.Reflection.TargetInvocationException: Exception has been thrown by the target of an invocation. ---> System.Exception: System.Runtime.Serialization.SerializationException: Type 'pwiz.Skyline.Model.LineColNumberedIoException' in Assembly 'Skyline, Version=20.1.0.155, Culture=neutral, PublicKeyToken=null' is not marked as serializable.
   at System.Runtime.Serialization.Formatters.Binary.WriteObjectInfo.InitSerialize(Object obj, ISurrogateSelector surrogateSelector, StreamingContext context, SerObjectInfoInit serObjectInfoInit, IFormatterConverter converter, ObjectWriter objectWriter, SerializationBinder binder)
   at System.Runtime.Serialization.Formatters.Binary.ObjectWriter.Write(WriteObjectInfo objectInfo, NameInfo memberNameInfo, NameInfo typeNameInfo)
   at System.Runtime.Serialization.Formatters.Binary.ObjectWriter.Serialize(Object graph, Header[] inHeaders, __BinaryWriter serWriter, Boolean fCheck)
   at System.Runtime.Serialization.Formatters.Binary.BinaryFormatter.Serialize(Stream serializationStream, Object graph, Header[] headers, Boolean fCheck)
   at SkylineTool.RemoteBase.SerializeObject(Object o) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\SkylineTool\RemoteBase.cs:line 60
   at SkylineTool.RemoteService.ProcessClientThread(Object streamArg) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\SkylineTool\RemoteService.cs:line 104
   --- End of inner exception stack trace ---
   at SkylineTool.RemoteClient.RemoteCallName(String methodName, Object[] arguments)
   at SkylineIntegration.Program.Main(String[] args) in c:\Users\Yuval\Documents\Skyline\3-27-17-tool\pwiz_tools\Skyline\Executables\Tools\PRMCalc\c#\SkylineIntegration\Program.cs:line 50

I am using as input:

0.0 0.0
NVVHQLSVTLEDLYNGATRKLALQK KLALQK 837.532@7,156.079@20 3103.692@1 2
GGKKGAVECCPNCR CKSCNGR 1018.4325@3,156.0786@4,57.0215@9,57.0215@10,57.0215@13    57.0215@1,1885.8485@2,57.0215@4    2
GGKKGAVECCPNCR ISPKDRCKSCNGR 1870.903@3,156.0786@4 156.0786@4,57.0215@7,1885.8485@8,57.0215@10 2
GGKKGAVECCPNCR KLALQKNVICDK 1665.9375@3,156.0786@4 156.0786@1,1885.8485@6,57.0215@10 2
GKNVVHQLSVTLEDLYNGATR CKSCNGR 1018.4325@2 57.0215@1,2451.2758@2,57.0215@4 2

Thank you!
Best wishes,
Tatiani.
view request
Instrument setting issue - Document grid bug
(3 responses) Tobi 2020-06-08

Dear Skyline team,

I noticed the Instrument settings/ Min Max m/z applies to both fragments and precursors at the same time. Did skyline always behave like this? I find it a bit troublesome as fragment measurement ranges are nearly all the time much wider than precursor measurement ranges. If I use this setting to properly filter precursors I loose lots of fragments, and if I properly filter fragments I spend alot of time on loading sometimes thousands of nonsense precursors into the target list.

Do you have a good tip or workaround on how I can differently filter precusors/fragments when building a target list?
Do you consider having separate settings for MS1 and MS2 in the near future? (just asking if by chance it makes sense for me to wait a bit).

Looking forward to a short reply.

[Skyilne-daily 20.1.1.158, add peptide PEPTIDER++, filter for 400 to 500 mz yields only 3 fragments]

Best wishes,
tobi

view request
Hide from View
(1 response) Jan Sklenar 2020-06-09

Hi Brendan,

I would like to be able to hide some chromatograms from view, without completelly removing them from document. Is it possible? The reason is, I like to keep all data for future purposes, but wish to compare only some runs in my QC data document.

Thanks,
Jan

view request
DIA-NN .speclib support
(17 responses) Tobi 2020-05-27

Dear Skyline Team,

could you please consider implementing support for DIA-NNs .speclib spectral libraries? Its a highly convenient tool for predicted libraries and much faster than Prosit.

https://github.com/vdemichev/DiaNN

With best regards,
tobi

view request
Shimadzu data file not loading
(3 responses) egauth 2020-06-08

Hi!
I'm encoutering a problem when loading a specific MRM Shimadzu data file inside Skyline 20.1.0.76. The error is:

Failed importing results file 'E:\Shimadzu Joe\20200601002_Repro_Cocaine_Hexane_003.lcd'.
The sample 20200601002_Repro_Cocaine_Hexane_003 contains no usable data.

I attach the Skyline method as well as the problematic .lcd data file. I also included another data file from the same MS but with a different MS method that loads correctly with the same Skyline method.

Can someone spot where the problem is?
Thanks!

 Integration_method.sky  DO_NOT_LOAD_20200601002_Repro_Cocaine_Hexane_003.lcd  LOAD_OK_20200129001_opt_012.lcd 
view request
Quantification of unlabeled peptides using SIL calibration curve
david schmidt 2020-06-08

Hi,

once again thanks for this great piece of software!

We are trying to quantify light (native) peptides using their isotope-labeled synthetic counterparts as standards. In contrast to the example in the "Absolute Quantification" tutorial, our calibration curve is generated with heavy peptides in a comparable matrix (and not with different light/heavy ratios). We do, however, spike a fixed concentration of heavy peptides into each sample to compensate for recovery differences.

The goal is to calculate the concentration of the light peptides using the labeled peptide's calibration curve and then to normalize that concentration value to the recovery (measured conc. heavy/spiked conc. heavy). My problem is, how to apply the heavy peptide-based calibration curve for the quantification of light peptides. Is there a way to specify the "internal standard type" separately for the calibration curve and the samples?

Thank you,

David

view request
His-tag peptides
(2 responses) jung165 2020-06-04

Hi Skyline team.
I always appreciate for the fascinating tool that is applicable for targeted protemics!

I have one quick question about selecting peptides.
I prepared the his-tag purified and enriched proteins which are transfected in Ecoli.
Therefore, I would like to insert modification about His-tag on my target peptide but, in modification tap I could not find such modification.
Is it applicable to just put 6H(histidine) after my target peptide and inset on Skyline??

I really appreciate for your help
Sincerely,

view request
Support for RDkit isotope aware
(3 responses) landerson 2020-05-26

Does Skyline support RDkit isotope aware molecular formulas? RDKit uses nomenclature that a heavy carbon is [C13]. For example, if 2 carbons in glucose were heavy you might use C4[C13]2H12O6.

I know of the apostrophe " ' " method e.g. C’C5H12O6 that is embedded in Skyline. The RDkit nomenclature feature would make the synergy between a postgresql database more streamlined.

view request
Peptide Uniqueness
(1 response) Hui 2020-06-02

Hello Skyline Team,

Hope all is well.

I am trying to create an SRM list where we have human samples with mouse heavy standard. Is there a function that works the opposite of "Exclude Background Proteome" under "Edit -> Unique Peptides..."? The goal is to choose peptides that are shared between both human and mice genomes.

Thank you!

view request
Fix for QuaSAR download error
(2 responses) jung165 2020-03-02

Hello, Skyline team

I have one question about downloading QuaSAR.
I am currently using Skyline 20.1.

When I try to download QuaSAR, it automatically downloads R 3.0 and continue to download QuaSAR related package.
However, the error message appealed with mentioning I need R 3.5 or higher version to download the packages.
So, I downloaded R 3.5 but, error message still appeals and it seems that those packages were still trying to be downloaded in R 3.0 folder.
May I ask your help for this issue??

I appreciate for your help

Skyline

The following packages failed to install:

bitops
reshape
gtools
ggplot2
gplots

view request
Import PRM results from Xevo QTof
(9 responses) taleb sedighi 2020-05-29
Dear Skyline Support Team, I have a problem in importing PRM result files from Masslynx (acquired on a Waters Xevo G2-XS QTof) into the Skyline. The PRM data includes one MS scans and 5 MSMS scans. Importing completes without any error but Skyline only shows the precursor chromatograms, not the product ions. Because it only reads the MS scan, not the MSMS ones. Here, I have attached the selected transition settings as a ppt file. In the full-scan tab, "Targeted" is selected as the acquisition method; so Skyline should expect multiple MSMS scans but it does not read any of them. I would appreciate your help! Taleb
 PRM_transition_setting.pptx 
view request
Boxcar support
(13 responses) DiegoB 2020-05-13

Hi Skyline team
I was wondering if Skyline support MS1 filtering on data acquired using Boxcar on the Exploris.
Thanks a lot
Diego

view request
Unable to Import Result Files due to multiple chromatograms
(3 responses) stephanie 2020-05-28

Hi,

I am trying to import results (from a byonic search) for full scan filtering. I created my library and a background proteome. I have a custom labeling system with labeled R/M and methyl groups. My samples are fractionated. I have tried adding samples as multiple replicates with multiple injections, single injection, and one new replicate but I get a similar error message (see below).

I was working with three libraries and importing multiple replicates. I am going to go back and re-build my library as a single library. Not sure if that will help.

Do you have any suggestions? If there is one peptide like this, there will be more.

Best,
Stephanie

Error Message:
At 8:44 PM:
Failed importing results file 'C:\Users\slynn\Documents\3Plex_Fraction_Skyline\ADMA Fraction\20200113_SML_ADMACombo_HILIC_10.raw'.
Unable to process chromatograms for the molecule 'YMHR[+18]NR' because one chromatogram ends at time '23.7843837738037' and the other ends at time '25.4307174682617'.

Inner exceptions:
Exception type: System.InvalidOperationException
Error message: Unable to process chromatograms for the molecule 'YMHR[+18]NR' because one chromatogram ends at time '23.7843837738037' and the other ends at time '25.4307174682617'.
Unable to process chromatograms for the molecule 'YMHR[+18]NR' because one chromatogram ends at time '23.7843837738037' and the other ends at time '25.4307174682617'.
at pwiz.Skyline.Model.Results.PeptideChromDataSets.AddDataSet(ChromDataSet chromDataSet) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Results\PeptideChromData.cs:line 838
at pwiz.Skyline.Model.Results.PeptideChromDataSets.Add(PeptideDocNode nodePep, ChromDataSet chromDataSet) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Results\PeptideChromData.cs:line 794
at pwiz.Skyline.Model.Results.ChromCacheBuilder.AddChromDataSet(Boolean isProcessedScans, ChromDataSet chromDataSet, PeptidePrecursorMz peptidePrecursorMz, IDictionary2 dictPeptideChromData, ICollection1 listChromData, ChromFileInfo fileInfo) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Results\ChromCacheBuilder.cs:line 670
at pwiz.Skyline.Model.Results.ChromCacheBuilder.CalcPeptideChromDataSets(ChromDataProvider provider, List1 listMzPrecursors, HashSet1 setInternalStandards) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Results\ChromCacheBuilder.cs:line 510
at pwiz.Skyline.Model.Results.ChromCacheBuilder.Read(ChromDataProvider provider) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Results\ChromCacheBuilder.cs:line 371
at pwiz.Skyline.Model.Results.ChromCacheBuilder.BuildCache() in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Results\ChromCacheBuilder.cs:line 252

view request
Selective peak integration based on idotp
(4 responses) Fabian 2018-05-29

Dear all,

i would like to have the option to integrate only peaks above a certain idotp threshold.
At the moment, the option [refine_advanced_results] Min idotp: decides if a peptide is kept or not.
I would like to have the same option based on the peaks not on the peptide.

Did I miss sth. or are there workarounds (peak training model did not convinced me).

Best regards

Fabian

view request
LOQ CV requirement iterates from low to high concentration, but high to low concentration would be better
(1 response) philip remes 2020-05-28

Hi guys,

I'm trying to use the Calibration Curve functionality, and have set the LOQ CV requirement to 20%. I notice some strange stuff going on, where sometimes a compound will have crap CV's at low concentration, then randomly have a level with a CV less than 20% when there's just noise. This level will get picked as the LOQ, even though some higher concentrations have worse CVs. Looking at the data, it's clear that the true LOQ should be at much higher concentration. To me, this looked as though the algorithm was proceeding from low concentration to high concentration, and taking the first level that meets the CV requirement as the LOQ. I would suggest that the LOQ would be more accurate if you proceeded from high concentration to low concentration, and picked the last level that meets the CV requirement. The peptide IFYNQQNHYDGSTGK is one example of this.

Thanks
Philip

view request
Moel Method for Scheduled PRM
z-proteomics 2020-05-28

Can any skyline experts with experience of using QE HF machine provide me a model PRM (Scheduled) method with detailed parameter setting?
Greatly appreciate your helpn in advance.
--- OmicSky

view request
unable to import HD-MRM result files into skyline in the last release version/daily version
(2 responses) simon daled 2020-05-28
i am unable to import my HD-MRM runs into skyline. It works fine in the 19.1 release version, but not in the newest version, there must be some bug somewhere. The runs are partially imported but the shown results are completely wrong.
Is it possible to download the previous version somewhere?
Thank you!
view request
Some small molecules don't support calibration curve
(1 response) philip remes 2020-05-27

Hi guys,

I have a small molecule document that is being used to analyze a dilution curve. Of 79 molecules, 2 of the calibration curves give the message "Error: All of the external standards are missing one or more peaks". This would appear to be similar to other reported issues.

Other support requests that mention this error, but don't seem to help me

Similar Issue 1
Similar Issue 2
Similar Issue 3

However, there is no internal standard normalization, the Analyze Concentrations are set, and the Sample Types are set to standard. Additionally it's kind of weird that just two of them do this, when they both have reasonable looking Peak Area plots vs Analyte Concentration.

Thanks
Philip

view request
method export error
(3 responses) dawn dufield 2020-05-22
Hi.

do you know what would cause this error. We have the analyst configuration in simulation mode and seems to be able to open the methods properly. This same skyline file can be exported on a different computer with analyst. So it feels like an analyst config issue, but seems like it should work.

any suggestions

---------------------------
Skyline
---------------------------
An error occurred attempting to export.
ERROR: Unable to cast object of type 'Sciex.MultiQuant.AnalystImprs.ApplicationObject' to type 'Interop.Analyst.ApplicationClass'.

Command-line: Method\AbSciex\TQ\BuildQTRAPMethod -s -m "X:\Analyst Data - BioPharma\Projects\Peptide\Acquisition Methods\Trap Pepcal 10 min.dam"
Working directory: X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods
Standard input:
X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods\~SK14E.tmp
X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods\test.dam
1752.439942,1930.069112,5,sequence1.xremoved this sequence info....
---------------------------
OK More Info
---------------------------
System.IO.IOException: ERROR: Unable to cast object of type 'Sciex.MultiQuant.AnalystImprs.ApplicationObject' to type 'Interop.Analyst.ApplicationClass'.

Command-line: Method\AbSciex\TQ\BuildQTRAPMethod -s -m "X:\Analyst Data - BioPharma\Projects\Peptide\Acquisition Methods\Trap Pepcal 10 min.dam"
Working directory: X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods
Standard input:
X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods\~SK14E.tmp
X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods\test.dam
1752.439942,1930.069112,5,sequence removed....

---> System.IO.IOException: ERROR: Unable to cast object of type 'Sciex.MultiQuant.AnalystImprs.ApplicationObject' to type 'Interop.Analyst.ApplicationClass'.

Command-line: Method\AbSciex\TQ\BuildQTRAPMethod -s -m "X:\Analyst Data - BioPharma\Projects\Peptide\Acquisition Methods\Trap Pepcal 10 min.dam"
Working directory: X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods
Standard input:
X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods\~SK14E.tmp
X:\Analyst Data - BioPharma\Projects\Ext\Acquisition Methods\test.dam
1752.439942,1930.069112,5,sequences removed.....

   at pwiz.Common.SystemUtil.ProcessRunner.Run(ProcessStartInfo psi, String stdin, IProgressMonitor progress, IProgressStatus& status, TextWriter writer) in C:\proj\skyline_20_1_x64\pwiz_tools\Shared\Common\SystemUtil\ProcessRunner.cs:line 62
   at pwiz.Common.SystemUtil.ProcessRunner.Run(ProcessStartInfo psi, String stdin, IProgressMonitor progress, IProgressStatus& status) in C:\proj\skyline_20_1_x64\pwiz_tools\Shared\Common\SystemUtil\ProcessRunner.cs:line 57
   at pwiz.Skyline.Util.Extensions.UtilProcess.RunProcess(ProcessStartInfo psi, String stdin, String messagePrefix, IProgressMonitor progress, IProgressStatus& status) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Util\Extensions\UtilProcess.cs:line 44
   at pwiz.Skyline.Model.MethodExporter.ExportMethod(String exeName, List`1 argv, String fileName, String templateName, Dictionary`2 dictTranLists, IProgressMonitor progressMonitor) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Export.cs:line 3906
   at pwiz.Skyline.Model.AbiMethodExporter.ExportMethod(String fileName, String templateName, IProgressMonitor progressMonitor) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Export.cs:line 2397
   at pwiz.Skyline.Model.ExportProperties.<>c__DisplayClass119_0.<ExportAbiQtrapMethod>b__0(IProgressMonitor m) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Export.cs:line 523
   at pwiz.Skyline.Util.ProgressWaitBroker.PerformWork(ILongWaitBroker broker) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Util\UtilUI.cs:line 125
   at pwiz.Skyline.Controls.LongWaitDlg.RunWork(Action`1 performWork) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Controls\LongWaitDlg.cs:line 246
   --- End of inner exception stack trace ---
   at pwiz.Skyline.Util.Helpers.WrapAndThrowException(Exception x) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Util\Util.cs:line 1907
   at pwiz.Skyline.Controls.LongWaitDlg.PerformWork(Control parent, Int32 delayMillis, Action`1 performWork) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Controls\LongWaitDlg.cs:line 194
   at pwiz.Skyline.Controls.LongWaitDlg.PerformWork(Control parent, Int32 delayMillis, Action`1 performWork) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Controls\LongWaitDlg.cs:line 131
   at pwiz.Skyline.FileUI.ExportDlgProperties.PerformLongExport(Action`1 performExport) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\FileUI\ExportMethodDlg.cs:line 2022
---------------------------
view request
Exporting user-defined reports from command line
(2 responses) fcsigloch 2020-05-27

Hi,
I am trying to export a custom report from command line. I defined the report previously out of the GUI and can access it from there. However, when I try to run
SkylineCmd.exe --in=someFile.sky --report-name="MyResults",
I get the error: Error: The report MyResults does not exist.

I tried to fix the problem by running SkylineCmd.exe --in=someFile.sky --save-settings, but that did not help.
(However save-setings seems to have fixed a similar problem with a user-defined mProphet model. This can now be used for reintegration.)

How do I correctly export the report?

Greetings, Florian

P.S.: Sorry for posting at the wrong place at first!

view request
Missing transitions in timsTOF data
(1 response) matt 2020-05-26

Hi

I have some timsTOF Pro PRM data which I am having problems with. For some of my targets Skyline does not find the MS2 fragments but I can clearly see them using Bruker DataAnalysis software. It only seems to be selected targets, some work fine.
Any suggestions?
Thanks!
Matt

view request
Pivot Editor Median Feature
(4 responses) roman sakson 2020-05-20

Dear Skyline Team,

first of all, thank you for adding the pivot editor capability to the report grids, making Skyline even more versatile! I was wondering whether it would make sense to add the option median to what the editor can do, as at the moment only the mean seems available. I think that the median could be helpful at least in some cases as it is more robust against outliers than the mean.

Thank you for consideration,

Roman

view request
How to use CIRT to predict retention time?
(3 responses) caixue 2020-05-20

Hi,
I am trying to use the peptides selected by myself as CiRT to predict the retention time. The irt value of each CiRT peptide has been calculated and saved as a new Retention time predictor. Please see the attachment. However, when I chose the predictor, the document could not import the cirt peptides I selected, so I could not import the result. What should I do?
Looking forward to a reply. Thanks!

 Skyline cirt problem.pdf 
view request
MaxQuant spectral library
(1 response) xzhong9 2020-05-20

Hi Skyline Team,
I would firstly like to thank you for enabling Skyline for DIA data analysis.
I have a question about using MaxQuant msms.txt file to construct a spectral library in Skyline. The two samples are labeled with two mass-defect tags with a mass difference of 18 mDa. To construct the spectral library, the mixed samples were acquired in DDA mode, searched in Proteome Discoverer or MaxQuant, and import the .msf or msms.txt file into Skyline. I want to compare the results difference between PD and MaxQuant. I am able to get great results using .msf from PD, but have no clue why Skyline is not able to resolve the light and heavy fragment ions using msms.txt from MaxQuant as shown in the attached files. I would appreciate it if you could help me resolve it.

Thank you,
Xiaofang

 PD.JPG  MaxQuant.JPG 
view request
Error parsing spectrum while building a library
(6 responses) pamrollahi 2020-05-18

Hello,
I am trying to build a new library using a ".msf" file exported from PD and get this error (attached snapshot).
I was wondering if anyone here has faced such an issue and can kindly help me solve it.
Thanks!
Pouya

 Capture.PNG 
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MSP Spectral Library import for small molecules failed
(2 responses) stolltho 2020-05-17
Hi there.

Wanted to import .msp spectral libraries from:
https://mona.fiehnlab.ucdavis.edu/downloads
https://minedatabase.mcs.anl.gov/#/download
http://prime.psc.riken.jp/compms/msdial/main.html#MSP

.msp import works for HMDB, MoNA and ReSpect but NOT for GNPS, Fiehn HILIC and LipidBlast (throws an error message, see example attached) from https://mona.fiehnlab.ucdavis.edu/downloads

Spectral library import from
https://minedatabase.mcs.anl.gov/#/download
http://prime.psc.riken.jp/compms/msdial/main.html#MSP
does not work at all, i.e. shows zero molecules after import

win10
skyline-daily 20.1.1.134

cheers,
Thomas
 lipidblast error.png 
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Importing Data
(7 responses) dawn dufield 2020-05-07

Hi I have 4 questions

  1. Is there a way to import acquired data that you don't have a skyline file for and have skyline automatically create transitions for those that are in the data file? vs creating it first in skyline and then importing. This would be much quickly for some small molecule data

  2. How does skyline handle Sciex Qtrap IDA data? If you acquire an IDA data file say for the top 4. Is there an easy way to review that data in skyline?

  3. When I'm integrating a specific peak and try to "apply to all" many times it does not still pick the same peak. How can I force this so I don't have to select each window in every file. (This is especially true when data is not great or peaks are noisy or low)

  4. What is the "best conditions" for transitions settings in your opinion for finding the best peptide to follow. Should you really cast a very wide window and use all transitions to last ion?

thanks
Dawn

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How does skyline store and handle the settings files
(1 response) hogan 2020-05-06

Hello Skyline Team,

Firstly I hope you and your families are well!

I have come across some strange behavior with trying to update and save the settings for a skyline document to be used elsewhere. How does skyline handle the .skys files. The only way I have ever seen a .skys file is by using the "Share.." command in the settings drop-down to generate one. Where do all the other .skys files live and how does skyline interact with them? Are they specific to a local installation of skyline? I can provide very detailed and specific examples if necessary.

Thanks!

Kyle

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Panorama error with Skyline daily document
(3 responses) Chris Ashwood 2020-05-14

Hi all,

I hope you are well. I have been encountering a reproducible error lately, working on a Skyline document that uses the new log iRT feature. There are other niche features used including a spectral library of 500 molecules. The document itself is sensitive and sizeable (95 MB) but I can share it privately. Please find the Panorama warnings/errors below:

14 May 2020 20:23:29,961 WARN : The version of this Skyline document is 20.11. This is newer than the highest supported version 4.
14 May 2020 20:24:06,450 INFO : 98% Done

14 May 2020 20:24:06,677 INFO : Done parsing Skyline document.

14 May 2020 20:24:10,809 INFO : Failed to complete task 'org.labkey.targetedms.pipeline.TargetedMSImportTask'

14 May 2020 20:24:10,812 ERROR: SqlExecutor.execute(); uncategorized SQLException for SQL []; SQL state [25P02]; error code [0]; ERROR: current transaction is aborted, commands ignored until end of transaction block; nested exception is org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block

Caused by: org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block

Caused by: org.postgresql.util.PSQLException: ERROR: duplicate key value violates unique constraint "uix_auditlogentry_document"

Detail: Key (documentguid, entryhash)=(577e6540-bd2e-4a40-85dd-c009a80ba168, VeCVOeCev0LQhxtTovC3nshjfOY=) already exists.

14 May 2020 20:24:10,853 ERROR: SqlExecutor.execute(); uncategorized SQLException for SQL []; SQL state [25P02]; error code [0]; ERROR: current transaction is aborted, commands ignored until end of transaction block; nested exception is org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block

org.springframework.jdbc.UncategorizedSQLException: SqlExecutor.execute(); uncategorized SQLException for SQL []; SQL state [25P02]; error code [0]; ERROR: current transaction is aborted, commands ignored until end of transaction block; nested exception is org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block

Caused by: org.postgresql.util.PSQLException: ERROR: current transaction is aborted, commands ignored until end of transaction block

Caused by: org.postgresql.util.PSQLException: ERROR: duplicate key value violates unique constraint "uix_auditlogentry_document"

Detail: Key (documentguid, entryhash)=(577e6540-bd2e-4a40-85dd-c009a80ba168, VeCVOeCev0LQhxtTovC3nshjfOY=) already exists.

Cheers,
Chris

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DIAPASEF Scoring Issues
(9 responses) Max McCabe 2020-05-12
I have been attempting to use Skyline for processing DIAPASEF data from analysis of neat plasma on the timsTOF Pro. The issue is that although everything during setup and upload appears to be proceeding successfully, I am unable to achieve separation between target and decoy distributions when generating a scoring model. Additionally, the data have a lot of interference in most of the ID chromatograms. I am unsure if the issues are due to library quality, Skyline import settings, or improper integration of ion mobility.

 I've set up the Skyline document as described in the Large-Scale DIA Webinar with a couple of alterations to accommodate ion mobility data where I have been able to find the settings. I'm using an ion mobility resolving power of 40 following recommendations in an earlier support thread and have tried both centroided (assuming that this process is internally supported) and TOF analyzer settings when uploading raw .d data with the same results. The DIA scheme is imported directly from the data and appears to be correct.

I am using a spectral library generated using MSFragger with a 0.99 score cutoff and have checked to confirm that the library contains ion mobility values as well as RT.

I've played around with the setting and am at a loss for what the issue could be because the data has been successfully searched using other pipelines. I have zipped the .sky and associated files into an uploaded folder titled "Skyline Support Upload".

Thanks in advance for the help!
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RT peptides
(12 responses) smanda 2020-04-14

Hi Brendan,

We have 50 peptides (including some from Biognosys) which we use as RT peptides in all samples. I was wondering if there is a way I can use all of them for my RT alignment in Skyline? I see currently it takes 25 or so and says 20 required. Am I missing something.

Regards,
Srikanth

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Only coeluting product ions as quantitative (extended)
(8 responses) Tobi 2018-12-17

Dear Skyline team,

I noticed that setting the "Quantitative" checkbox also works with copy and paste, but no matter if done by copy and paste or by manual clicking, it seems impossible to get the same checkboxes as in the Coeluting column because of constant recalculations overwriting manually set checkboxes (in PRM)

For just a single file one could think of exporting and reimporting a transition list filtered for coeluting product ions, but for a larger experiment, this is not feasible. Is it possible to somehow fix the quantitative column to make it directly susceptible to manual editing (in general just setting "not coeluting" to "not quantitative")? Is there a smarter way to do that?

I know about unchecking integrate all, but it would be much better to have full control over the quantitative state of a transition compared to just removing the target from the target list which impacts all datasets.

The attached file shows what would be the desired output, its just not fully implementable for a whole experiment.

Thank you for all thoughts and ideas on this topic.

Best,
tobi

 181116_PRM_quant_coeluting.sky.zip 
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Using Prosit for heavy ions
(7 responses) denisoleynik3007 2020-05-08

Does Prosit work for a prediction of MS/MS spectra of heavy ions (in particular heavy y-ions)?

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About Synapt G2-Si (Waters). Data-independent acquisition
(3 responses) eriosc 2020-05-05

Hello!

I would like to know a few things about Skyline.

I have used commercial software most of the time; however I would like to know if Skyline supports * .raw files from Waters coming from a Synapt G2-Si mass spectrometer. I ask this because I am interested in doing Label-free (DIA) using HDMSE and UDMSE mode. Is it possible to do this?

Additionally, I would like to know if with Skyline it is possible to identify crosslinked peptides (XL-MS) using the same way HDMSE and UDMSE (DIA). If possible, it would be great because I have some ideas to develop using this methodology.

Thanks and regards

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issue when importing peptide search from a MaxQuant msms.txt file
(5 responses) marie locard-paulet21061 2020-05-01

Hello, I am trying to import the results of a DDA search from MaxQuant (1.6.14) for generating a library in Skyline (for DIA data analysis).
I work with Skyline (64-bit) 20.1.0.76 (0d62150d0). My samples do not contain iRTs, and kept all default parameters for the "import peptide search".

Here is the error message that I get (copied and pasted from Skyline):

---------------------------
Skyline
---------------------------
ERROR: No spectra were found for the new library.

Command-line: C:\Users\malopa\AppData\Local\Apps\2.0\5ZNXQLMA.P6A\TRMG87OZ.R8T\skyl..tion_e4141a2a22107248_0014.0001_72292abfbc2fefb5\BlibBuild -s -A -H -o -c 0.95 -i MQDDALib_MC1_Carba -S "C:\Users\malopa\AppData\Local\Temp\2\tmp3F62.tmp" "C:\data\Skyline\MQDDALib_MC1_Carba\MQDDALib_MC1_Carba.redundant.blib"
Working directory: C:\data\MQ\HeLA_21min_Library\MQDDA_MC1_Carba\combined\txt
---------------------------
OK More Info
---------------------------
System.IO.IOException: ERROR: No spectra were found for the new library.

Command-line: C:\Users\malopa\AppData\Local\Apps\2.0\5ZNXQLMA.P6A\TRMG87OZ.R8T\skyl..tion_e4141a2a22107248_0014.0001_72292abfbc2fefb5\BlibBuild -s -A -H -o -c 0.95 -i MQDDALib_MC1_Carba -S "C:\Users\malopa\AppData\Local\Temp\2\tmp3F62.tmp" "C:\data\Skyline\MQDDALib_MC1_Carba\MQDDALib_MC1_Carba.redundant.blib"
Working directory: C:\data\MQ\HeLA_21min_Library\MQDDA_MC1_Carba\combined\txt
   at pwiz.Common.SystemUtil.ProcessRunner.Run(ProcessStartInfo psi, String stdin, IProgressMonitor progress, IProgressStatus& status, TextWriter writer) in C:\proj\skyline_20_1_x64\pwiz_tools\Shared\Common\SystemUtil\ProcessRunner.cs:line 62
   at pwiz.BiblioSpec.BlibBuild.BuildLibrary(LibraryBuildAction libraryBuildAction, IProgressMonitor progressMonitor, IProgressStatus& status, String& commandArgs, String& messageLog, String[]& ambiguous) in C:\proj\skyline_20_1_x64\pwiz_tools\Shared\BiblioSpec\BlibBuild.cs:line 201
   at pwiz.Skyline.Model.Lib.BiblioSpecLiteBuilder.BuildLibrary(IProgressMonitor progress) in C:\proj\skyline_20_1_x64\pwiz_tools\Skyline\Model\Lib\BiblioSpecLiteBuilder.cs:line 160
---------------------------


I attach to this post the msm.txt files and corresponding mqpar.xm.
please let me know if you need more information.

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Setting up Skyline in a Virtual Environment
(2 responses) halve021 2020-05-01
The University of Minnesota is working toward a virtual delivery pilot in the research space. Can you tell me if the license for Skyline allows the software to run in a Citrix or XenApp environment?

If this is not possible, can they be delivered via Remote Desktop Services?

Thank you for your time,
...Justin Halverson
IT - University of Minnesota
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Refinement Suggestion: Min peptides per protein EXCEPT X
alejandro.cohen 2020-05-01

Hi Skyline people

Suggestion for refinement in Skyline\Advanced menu:

Add an option to remove Min peptides per protein with another tick box reading 'EXCEPT for proteins smaller than': X (Da). The value of the intact protein could be estimated from the FASTA file, maybe?

Some smaller proteins (<5-10KDa) give very few good (sometimes only one!) unique tryptic peptides, yet very good quality and signal. You dont want to discard them just because you didn't get enough number of peptides.

Your thoughts?

Alex

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Manual integration of SST is not transferred to AutoQC/LabKey
(3 responses) katrin freiburghaus 2020-04-27

Dear Skyline team

I am using Skyline with AutoQC Loader to get my SST uploaded to the Panorama data management system, where I can track the instrument performance (Xevo TQ-S triple quadrupole, Waters). All is working nicely, excpet when the Skyline algorithm picks a wrong peak in my data. I can adjust it manually in the Skyline file, but it is not updated in Panorama.

Could you tell me what I need to do so that my manual adjustments are adapted in Panorama?

Thank you very much and kind regards,
Katrin

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General question about peptide quantitation
(2 responses) caho 2020-04-29

Hi there,

May I please ask, with regards to peptide quantitation, is the sum of the peak areas of all fragment ions taken (so the total peak area of the precursor, precursor [m+1], precursor [m+2] fragment ions), or is generally the fragment ion with the greatest peak area used? May I also please ask how is the peak rank determined for each peptide/fragment ion?

Thank you so much for your help!
Cally

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Precursor metabolite data from SWATH
(3 responses) lefelit 2020-04-09

Hi

I have metabolite data from TOF MS acquisition on a SCIEX triple TOF instrument and I am trying to extract the ion chromatogram information for more than 200 precursors. I am not interested at this stage for fragment data. I built a method with all the molecules of interest with the accurate masses and when i applied it to a test mixture of pure standards the software does not find almost 60% of them. I set the mass window to 0.2 and the RT window to 1min and still does not find them. What could be the source of the problem? How could I also customise the threshold setting to different value per precursor?

Many thanks

view request
Installing Slyline error
(4 responses) alejandro.cohen 2020-04-28

Hi Skyline people,

I moved to home office, and I'm trying to install Skyline (regular or daily) on my laptop running Window10 , and in both cases I get the following error:

An error occurred trying to download 'https://skyline.gs.washington.edu/software/Skyline-release-64_20_1/Skyline.application'.
See the setup log file located at 'C:\Users\Agus\AppData\Local\Temp\VSDF497.tmp\install.log' for more information.

BTW, I can't seem to find that Temp folder in the C: drive anywhere...

Any ideas?

Thanks

Alejandro

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MSP Spectral Libraries for Small Molecules
(6 responses) Christina lucas 2020-04-22

Hi there,

i am trying to Import an MSP library to Skyline.

First attempt was an Test Library created in ThermoFishers mzVault exported as MSP (attached). Which was not working, no element was shown.
Second attempt, as Thermo might have some Export issues on that was to download the HMDB (or any other file in MSP) from https://mona.fiehnlab.ucdavis.edu/downloads

Still I get no content in my library.

So far I could not find anything in Forum thats giving me a hint how to open them correctly.
I am using daily 20.1.1.83 on a Win 10.

Best,

Christina

 Acyl_Test.msp 
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Prosit .msp library import is missing iRT values
(4 responses) anatoly.urisman 2020-04-25

Hi Skyline Team,
I was wondering if the iRT values in Prosit generated .msp libraries could be parsed upon import into Skyline. I use a simple conversion script as a work-around, but this adds an extra step. I suppose another solution would be to change Prosit output to Skyline-readable.
Thanks!
Anatoly

Here is an example of Prosit .msp format...

Name: TIFGGSDSTNQGPSNGER/2
MW: 912.41136795554
Comment: Parent=912.41136795554 Collision_energy=28.0 Mods=0 ModString= TIFGGSDSTNQGPSNGER///2 iRT=32.720001220703125
Num peaks: 46
175.118952167 0.10104765714257913 "y1/0.0ppm"

view request
iRT-Pierce combo library import converts all unmodified peptides to K+8, R+10
(1 response) anatoly.urisman 2020-04-14

Hi Skyline team,
I really like the feature where you can combine multiple blib libraries in Import Peptide Search wizard to create a new combo library. This allows me to use very different search results for iRT peptides (I use Peirce iRT mix, which is K+8 and R+10 modified) and for target peptides (which are usually unmodified) to generate blib files.
However, when I use this combo library to add target peptides, unmodified peptides are incorrectly showing as K+8 and R+10 modified (even though the reference sequences and spectra are correctly showing as unmodified in the library browser). Decoys are also modified as a result. As a work-around I can remove the modifications using Refine --> Advanced --> Remove Label Type (heavy), but I have a feeling that this was not the intended behavior during target import, so just wanted to bring it to your attention.
I've uploaded a file that illustrates this problem (DIA_2020-03-14_90min_v2.sky.zip).
Thank you for looking into this!
Anatoly

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noenzyme setting
(1 response) egulyas 2020-04-27

Dear Brendan,

I would like to do peptidomics study. The identified peptides have no specificity on any side. How can I set it in the "digestion" ? (both side unspecific or no enzyme)
Thanks,

Eva

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A question about MSstats
(4 responses) 1214800593 2020-04-26

Hi
I want to do a DIA project, but when I use the MSstats,it report error like attachment.
please help me!
THANKS!
Steven Liu

 MSstats.png 
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pepxml file generate
(3 responses) pucci biagio 2020-04-23

Hi,
i'm working with bruker tof and I'm use mascot as engine software...I'm not able to generate pepxml file to use for MS1 DDA analysis..
help me!!
regard

view request
Skyline is CFR part 11 compliant software
(1 response) Stephane CHARMONT 2020-04-23

Hi,

I'm currently working on pharma environment and specially at quality Level 2. But we are doing business with external service provider for clinical trials analysis where their lab need to be GxP qualified.
I would like to know if Skyline software is CFR part 11 compliant software and at the same time I would like to know if skyline incorporate an Audit trail system as usual qualified software from MS supplier or vendor?

Thanks a lot

Regards

Steph

view request
Peak selection for MRM
(1 response) bart van puyvelde 2020-04-24

Hi Brendan and Skyline team,

I am currently doing some data analysis for an MRM experiment and I got a question concerning the peak selection.

I would like to force Skyline to always select the peak within a 0.1minute RT window or even smaller.
I tried to do this by changing the Full-Scan settings, but apparently it does not work.

Any suggestions to get this working? I can't share the Skyline file through the support page because it contains patient data.

Best,

Bart

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Peak with high mass error not found
(2 responses) katrin freiburghaus 2020-04-23

Dear Skyline team

I encountered the following problem: Our Q-Exactive Orbitrap was not well calibrated and produced data with high mass errors (up to 70 ppm deviation) for all small molecules. I imported the SST files into Skyline and it did not integrate any peaks (see attachement "Skyline_Kynurenic_acid") using the attached transition settings. But when I integrated the exact same file using TraceFinder software, I saw the peaks (attached TraceFinder screenshot for kynurenic acid, mass error -53 ppm).

I adapted the "method match tolerance m/z" in the Transition Settings to the allowed 0.6 m/z, but still no integration of my peaks.

Could you let me know what I am doing wrong and how I can get my results in spite of the high mass error?

Thank you very much for your help and best wishes
Katrin

 Skyline_Kynurenic_acid.PNG  TraceFinder_Kynurenic_acid.PNG  Skyline_Transition_Settings.PNG  Skyline_Transition_Settings_2.PNG 
view request
Unknown modifications
(3 responses) Wilfred Tang 2014-12-06
In building a library (Settings menu, select Peptide Settings, go to Library tab, click on Build button), it appears that Skyline does not like "unknown" modifications (modifications not mappable to Unimod(?) based on delta mass). See attached screen capture. I am using Skyline-daily (64-bit) 2.6.1.6899.

Why does Skyline need to know the modification name? Shouldn't the delta mass alone be sufficient information?

Is there a way to use "unknown" modifications in Skyline? We commonly deal with data having lots of glycan modifications - there are a wide variety of glycan modifications, and most are not in Unimod.

Thanks,
Wilfred
 Screenshot 2014-12-06 22.26.51.png 
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Pressure output?
(9 responses) alejandro.cohen 2020-02-20

Hi Skyline people,

I work with an Agilent 1290 coupled to a Qtrap5500 running MRM metabolomics which I analyze using Skyline (.dam methods, .wiff datafiles). Analyst is recording the column pressure output signals, however viewing these using Analyst is cumbersome and time intensive when working with big sample batches. We have encountered erratic behaviour of our chromatography, accompanied by unusual pressure profiles for samples, which tend to correct themselves after a few runs.

As part of our QC, it would be great if skyline would be able to add (overlap) the pressure traces on each chromatogram. Is this possible? I know each vendor 'hides' the pressure traces in different output and channels... but the information is there :)

I know this request has been done in the past... here another kind 'reminder'

Cheers

Alex

Alejandro Cohen, Ph.D.

Scientific Director
Biological Mass Spectrometry Core Facility. Room N-105
Life Sciences Research Institute
https://medicine.dal.ca/research/biological-mass-spectrometry.html

view request
FAIMS unit on Thermo Lumos
(8 responses) erik.soderblom 2019-11-20

Hi Skyline Team! I wanted to "bump" a thread entitled “Optimizing FAIMS on new Thermo instruments” from Martijn van Duijn back in September. We are now in the same boat (acquired a shiny new FAIMS device for our Lumos!) and at this point we just want to incorporate the data (say, 3 different CVs across 7 different peptides) into Skyline from our normal system suitability PRM acquisitions. I see where compensation voltage is a selectable unit under the ion mobility settings, but it doesn't look like Skyline is able to extract data for a single ("best") CV in the chromatogram view. Is this possible with current Skyline Daily (19.1.1.309) yet?
Thx,
Erik

view request
Exporting Isolation Lists
(5 responses) cjeline1 2016-01-29
Dear all, we are experiencing a similar problem as that documented by Josua Troesch on the 27th. Running skyline version 3.5.0.9319, exporting a Thermo fusion isolation list from a skyline file fails to render the generation of a .csv file. Do you have any suggestions to resolve this issue?

Thanks,

Pandey Laboratory
Johns Hopkins School of Medicine
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Set a Fixed Mass Tolerance in Skyline
(1 response) lancia darville 2020-04-20

We are processing small molecule data using Skyline and would like to know if there is a way to set a mass tolerance in Skyline? We have been observing a either a spilt or a broad peak for a targeted molecule when processing the data in Skyline which aligns with our stable isotope standard, however the mass error is ~10 ppm. If the same data in processed in Thermo’s XCalibur with a mass tolerance set to 5 ppm, the interfering peak is removed and we only see our target with a mass error between 0.5 and 1 ppm in the samples. We would like to be able to use Skyline to process all the data, however this will require us to set a specific mass tolerance. Please advise if a specific mass tolerance can be set in Skyline and if so how?

view request
Removing precursors for DIA data
(4 responses) epcasavant 2020-04-17

Hi,

I am trying to run my data through MS Stats and ran into the error that said I needed to remove the precursors. I saw your answer here (https://skyline.ms/announcements/home/support/thread.view?rowId=37139) and followed the following steps you outlined:

  1. View > Document Grid
  2. Choose "Transitions" from the Views dropdown at the top of the Document Grid
  3. Right click on the "Fragment Ion" column and choose "Filter"
  4. Set the filter type to "Starts With" and type "precursor" into the textbox.

However, how do I delete the precursors? When I select them all int he document grid, it looks like it literally deletes all of my data.

Thank you,
Ellen

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