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    Original URL path: https://skyline.gs.washington.edu/labkey/ (2014-08-11)
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    to employ cutting edge technologies for creating and iteratively refining targeted methods for large scale proteomics studies Build Targeted Methods Use prior knowledge of your proteins and peptides of interest to improve your measurement selectivity and precision See Quantitative Results Learn more about your quantitative measurements through the rich and interactive Skyline graphical data displays Store and Share Experiments Panorama stores the entire Skyline data model where you and your collaborators can explore it with a web browser Build Chromatogram Libraries Use Panorama to build fully validated chromatogram assay libraries you can use in Skyline for new experiments Project Page Support Panorama is a freely available open source targeted proteomics website knowledgebase for sharing experiments and validated assays that integrates into a Skyline proteomics workflow It can be installed on a local server or you can request a project on the PanoramaWeb org server hosted by the MacCoss Lab at the University of Washington Access privileges within a project may be customized allowing you to control fully who has access to data you publish to Panorama Project Page Support Topograph is a freely available open source Windows client application designed to analyze data in protein turnover experiments using chromatographic peak

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    to employ cutting edge technologies for creating and iteratively refining targeted methods for large scale proteomics studies Build Targeted Methods Use prior knowledge of your proteins and peptides of interest to improve your measurement selectivity and precision See Quantitative Results Learn more about your quantitative measurements through the rich and interactive Skyline graphical data displays Store and Share Experiments Panorama stores the entire Skyline data model where you and your collaborators can explore it with a web browser Build Chromatogram Libraries Use Panorama to build fully validated chromatogram assay libraries you can use in Skyline for new experiments Project Page Support Panorama is a freely available open source targeted proteomics website knowledgebase for sharing experiments and validated assays that integrates into a Skyline proteomics workflow It can be installed on a local server or you can request a project on the PanoramaWeb org server hosted by the MacCoss Lab at the University of Washington Access privileges within a project may be customized allowing you to control fully who has access to data you publish to Panorama Project Page Support Topograph is a freely available open source Windows client application designed to analyze data in protein turnover experiments using chromatographic peak

    Original URL path: https://skyline.gs.washington.edu/labkey/project/home/begin.view? (2014-08-11)
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    PRBB Barcelona Sep 28 Oct 3 2014 Winter Targeted Proteomics Workshop at IIT Bombay Dec 10 11 2014 Past Summer Skyline User Group Meeting at ASMS 2014 review the presentations Summer ASMS 2 Day Courses in Baltimore MD June 14 15 2014 Spring US HUPO Short Course in Seattle WA April 6 2014 Spring Targeted Quantitative Proteomics Course at UW Seattle March 31 April 4 2014 Spring ABRF Workshop in Albequerque NM March 22 2014 Winter SRM Course at ETH Zurich Feb 10 14 2014 review the tutorials Panorama Webinar II August 8 2013 watch the recording Summer SRM Course at ETH Zurich July 15 19 2013 review the tutorials Skyline User Group Meeting at ASMS 2013 review the presentations Quantitative Proteomics Course at US HUPO March 10 2013 review the workshop materials Panorama Webinar February 9 2013 watch the recording Skyline User Group Meeting at ASMS 2012 review the presentations Review the presentations Skyline User Group Meeting 2014 Recently Published Broudy Killeen Bioinformatics A framework for installable external tools in Skyline abstract Choi Bioinformatics MSstats an R package for statistical analysis of quantitative mass spectrometry based proteomic experiments abstract Egertson Nature Methods Multiplexed MS MS for improved data independent acquisition abstract Abbatiello Mollecular Cellular Proteomics Design Implementation and Multi Site Evaluation of a System Suitability Protocol for the Quantitative Assessment of Instrument Performance in LC MRM MS abstract Pages Skyline Targeted Proteomics Environment Install 32 bit Skyline v2 5 Install 32 bit Skyline v2 5 Unplugged Install 64 bit Skyline v2 5 Install 64 bit Skyline v2 5 Unplugged Release Notes Videos Video 1 Method Editing Video 2 Results Analysis Video 3 Existing Quantitative Experiments Video Trailer Tutorials 教程 中国语文 靶向 方法编辑 靶向方法优化 处理已有定量实验数据 MS1 全扫描筛选 靶向 MS MS PRM iRT 保留时间预测 チュトリアル 日本語版 ターゲットメソッドの編集 ターゲットメソッドの最適化 既存データ処理および定量実験 MS1フルスキャンフィルタ ターゲットMS

    Original URL path: https://skyline.gs.washington.edu/labkey/project/home/software/Skyline/begin.view (2014-08-11)
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    06 25 more less view request Building libraries from Glycopeptides 1 response john leszyk 2014 07 15 04 52 Hi Brendan and Kaipo Is it possible to build a library from spectra that contain glyco fragments with few to no peptide fragments How would one go about extracting the glyco fragments and using these to quantitate I have a glyco peptide that I can see a spectra in the raw file but it is dominated by fragments of the carbohydrate and un identifiable in a basic peptide search that includes the carbohydrate modification Thanks John more less view request MSF file library build error message 1 response john leszyk 2014 07 14 05 30 Hi Brendan I am getting this SQL build error see attached when trying to build from PD msf files Please advise Thanks John more less MSF file Library build problems pptx view request Data independent acquisition results 5 responses Alex Zhu 2014 07 09 07 14 Hi Brendan I am told skyline is able to import All Ions data with multiple CEs I am just wondering how skyline handles that results For example assume I used 4 CEs and apparently they have different product ion abundances how skyline then ranks the abundance of them Does it rank the average of the same product ion from 4 channels or does it use the result from the most abundant channel Thanks a lot Alex more less view request PRM error 3 responses xi 2014 07 09 05 52 Dear Brendan and Skyline Groups when I run PRM on Q exactive and analysis on the Skyline some peptides did not quantified There are no green yellow or red signals Are those peptides out of the detecion see attachment many thanks Best regards Xi more less skyline png view request Orbitrap Fusion multiplexed tMS2 question 2 responses craig braun 2014 07 08 15 16 Hi I m seeing a discrepency between the ratios that Skyline is reporting for a given peak in my file to what I can manually determine in xcalibur Attached is the Skyline file as well as the RAW file You can see that for the peak eluting at 9 2 minutes Light 649 37 heavy 659 38 the ratio between the precursors for example is about 1 2 whereas the ratio calculated in Skyline is approximately 1 1 I believe it mostly has to do with the weird peak shape that Skyline sees which is not the case in the RAW file I tried relaxing the mass accuracy as this varies slightly throughout the variation of the peak but this did not help Also I ve been running experiments like this multiplexed tMS2 that also have a multiplexed tSIM prior to the MS2 scan also on the Fusion but I can t seem to figure out a way to get skyline to load the MS1 spectra from these files Any help would be appreciated more less tMS2 Test sky view request format of peptide ID in pepXML file 3 responses Hongbo 2014 07 07 14 46 Hello Brendan I came across a problem importing pepXML files into Skyline We are in the transition of bioinformatic pipeline so there will be some difference in the pepXML files generated from the new pipeline pepXML files generated by the old pipeline worked great with skyline from which Skyline is able to get all the peptides from it But for the new one there are about 1 3 peptides missing in the Skyline library after importing pepXML files I attached one file that contains ID information of one peptide that was successfully imported by Skyline from the pepXML generated from the old pipeline but not from the new pipeline Can you check it to see which part of the information is missing or not recognized by Skyline so that I can ask out informatics people to change our new pipeline Let me know if you need more examples or information Thank you very much Hongbo more less pepXML from new and old pipeline docx view request Background Proteome issues 2 responses jrumsey 2014 07 02 09 44 I am concerned whether the unique peptide form is working properly As a check I added 2 proteins to my target list in skyline that are in my background proteome Shouldn t it find that all these peptides are shared with a protein in background proteome It does not do this For one protein it finds some shared peptides and for the other protein it does not find any Is there something else I need to do to determine if this is working appropriately more less view request proxy support 1 response ewindha 2014 07 01 13 03 Is there a way to enable proxy authentication support in version 2 5 64 bit If not do you plan to support this in the future or should people just use the offline version more less view request Retention time predictor jrumsey 2014 07 01 07 03 I am having a problem using the retention time predictor When I go to edit current ret time predictor or add a new one it will automatically choose a calculator used previously not the one I want I just want to use the SSRCALC and create a predictor from results run with my current gradient conditions When I click use results it automatically goes to a different calculator and selects peptides How is it selecting these peptides and how can I select different peptides to use with the calculator See attached file for screen shot more less ret time predictor 01July2014 pptx view request Results import 2 responses sstoychev 2014 06 30 10 34 Hi Brendan When importing MRM results I only see options for replicate analysis i e software treats all imported runs as replicates However what if one wants to import non replicates i e different concentrations treatment days etc Stoyan more less view request Can Skyline process AIF MS MS data 5 responses Prahlad Rao 2014 06 27 13 06 Hello I have collected protein spectral data on a Q Exactive The MS2 data has been collected using All ion fragmentation This is on a single protein I imported the files in to Skyline however Skyline only extracts MS1 spectra and absolutely no MS2 spectra Since this is the first time I have used AIF on a QE I obtained RAW files from Matthias Mann s 2010 paper on AIF http www ncbi nlm nih gov pmc articles PMC2953918 and tested them using Skyline whether it can extract MS2 data from those files and still Skyline is not able to extract MS2 spectral information Do I have to maybe change certain parameters in Skyline for it to properly read AIF data Thanks Prahlad more less view request crosslinked peptides donna 2014 06 27 07 09 Hi General question is it possible to view crosslinked peptides in Skyline So far I ve been able to see peptides which have the crosslinking reagent as a mass modification both light and heavy isotope labeled but none of the crosslinked peptides seem to be visible whether I try to import the spectral library from txt or pepXML mzXML Thanks donna more less view request export native instrument method files from Skyline to Waters Xevo 1 response fangmin xu 2014 06 26 17 53 Hello Brendan As the title addressed I was trying to export method file from Skyline to Xevo to run MRM CE optimization And in Skyline from File Export Method I chose Xevo and optimization with CE But I m not sure what format I should upload to the template file drop off window So I randomly uploaded one of my Xevo MassLynx MRMR method in an exp file but then after I licked Export Skyline gave me a error message saying The MassLynx is not correctly installed and the library quantifyClassLibrary can not be found The error message and the export setting window screenshots were attached here So my question is what file format I should put for this template file and what is the library mentioned here Thank you very much for your help Sincerely Angela more less Capture Error Skyline 20140626 PNG Skyline Export setting 20140626 PNG view request Multiple Peptide XIC bug 1 response Mike S 2014 06 26 13 03 In one of my documents at least one peptide is missing from the displayed XICs I noticed it because it has one of the highest intensities and yet is not shown I m using Skyline 2 5 0 6079 on a Win7 64bit computer I can email the document to whoever is assigned to the bug Mike more less view request SProCoP 2 responses Wilf 2014 06 25 03 52 Hi Brendan I think I am doing something silly I am wanting to use SProCoP and keep getting the following error message Error in scan file what nmax sep dec quote skip nlines na strings line 285 did not have 8 elements Calls tryCatch tryCatchList parse cmdline read table scan Execution halted Finished Could you help please All the best Andrew more less view request custom low mass reporters 1 response madeleih4709 2014 06 24 19 05 Hi Brendan Within Skyline is it possible to quant low mass reporters that are completely custom and derived from a large side chain modification Thanks in advance for your assistance Kind regards Madi more less view request LTQ data for label free quant 2 responses galeva 2014 06 24 10 57 We have raw data acquired on the LTQ mass spectrometer that we hoped to use for measuring MS1 peptide ion intensities The experiment was set up so MS1 low resolution spectra MS MS fragmentation spectra and ZoomScan for charge state determination spectra were collected While importing these raw files into Skyline we get an error message failed to build a cache error parsing ITMS p ESI d u Z ms bad lexical cast source type value could not be interpreted as target We would appreciate any ideas on how to enable Skyline to analyze this kind if data more less view request Importing peptides with a known charge state 3 responses Alon Savidor 2014 06 23 23 49 Hi Brendan Is there a way to import a list of peptides with their known charge state such that after the import Skyline will show only that one m z value per peptide instead of all possible m z s for each peptide as defined in the transition setting Thanks Alon more less view request CE field setting in LTQ orbitrap 1 response b104094039 2014 06 23 20 28 Hi Although the LTQ Orbitrap did not require CE field But I still need to select a option in CE field Which one is best Yu Chia more less Thermo LTQ Orbitrap Skyline Setting Problem ppt view request identifying unique peptides 5 responses jrumsey 2014 06 23 09 11 I am trying to come up with a list of proteotypic peptides I have a list of target proteins loaded in skyine which I performed an in silico trypsin digest I also have a background proteome loaded in skyline It is taking a long time to go through each target protein and then view unique peptides and deselect peptides in common with other proteins to get my final list of proteotypic peptides Are there any suggestions to do this in a more automated fashion Once selected I then need to be able to export a csv of these unique peptides and the protein they belong to more less view request How do I update Skyline daily to newest version 1 response mh167 2014 06 23 08 38 Hi the software was trying to update automatically when I restart it twice yesterday I decided to update it later since I was in a hurry However it won t update now even I restart many times I was trying to follow the link attached in the skyline daily notification email but I couldn t find the place to download the Skyline daily Current version is Skyline daily v2 5 1 5963 Thanks a lot in advance for your help more less view request Filtering product ions 1 response delphine pflieger 2014 06 18 01 41 Dear Brendan I am using Skyline to interpret PRM data and would like to understand the logic hidden behind the filtering process of the product ions in Transition Settings Filter what is the actual definition of the interval delimited by From and to Thank you in advance Best regards Delphine more less view request 15N label 2 responses pegah jalili 2014 06 17 15 35 Hi Brendan I want to find the incorporation of 15N labeling using skyline I set the isotope enrichment of Nitrogen to different value 95 to 99 Then I compare experimental result with expected monoisotpic M 1 M M 1 M 2 and M 3 using Full MS 1 I have several questions 1 How expected monoisotopic peaks were calculated did you consider abundance of H C N O S 2 Should I add abundance of each atom to the Isotope labeling enrichment table in transition setting then Sulfur is not there or if I am looking for N should leave other atoms empty 3 Do you have any tutorial or any paper that use Skyline for calculation of 15N enrichment 4 There is a article about a binary exe perl program from MacCoss lab can we accesses the program trough skyline site and if not skyline calculate the distribution the same ref for a binary exe perl program Measurement of the isotope Anal Chem 2005 77 7646 7653 Thank you more less view request Skyline skyline daily compatibility 1 response ronsein 2014 06 17 08 27 Hi I am using skyline daily but the MS instruments have the skyline I can t open my daily files on skyline Error the file contains an error on column 2 line 2 the document format version 1 7 is not supported Is there a way to make them talk Thanks Grazi more less view request How to fit collision energy CE setting 1 response b104094039 2014 06 15 08 45 Hi I use LTQ Orbitrap How to fix collision energy CE setting when i use LTQ Orbitrap XL Is it fit collision energy CE setting Thermo TSQ Vantage and Thermo TSQ Ultra in Skyline You Chia more less view request Charge reduced loss fragment 2 responses christopher loessner 2014 06 13 02 32 I am interested in a fragment that is originating from a chemical modification and resulting in charge reduction as well as a partial mass loss of this tag E g I have a m z 500 3 ion peptide plus tag that is resulting in a m z 700 2 fragment peptide minus a part of the tag as well as reduced charge state I have tried for some while to come to a solution but wasn t successful yet Has somebody an idea how I can annotate and by this quantify these ions Best regards Christopher more less view request Skyline for peptidomics work option for No Enzyme 1 response Jennifer 2014 06 12 14 46 We are currently using Skyline for peptidomic work We were trying to add or create a No Enzyme option in the list of proteases in the digestion tab settings because some of the peptide hits coming from our Mascot search results are endogenous peptides which were identified by Mascot when searched under No Enzyme and these endogenous peptides specific cleavages are not known However when we were trying to use Skyline we could not find a No Enzyme option so we decided to create one and add to the list but we had trouble creating it because it prompted us to provide amino acids to cleave from N or C terminal but it is difficult to do this because it is not as precise as how e g trypsin will do it like always on K R on C term May I ask if there is a way to work this out so we can efficiently use Skyline for peptidomics Hope to hear from you Thank you Jennifer Aguilan and Cristina Clement more less view request error message for Msstats QC in Skyline 1 response xi 2014 06 11 05 15 Hi I m new to MS stats and am trying to run a Msstats QC from skyline to see how it works I m running MS stats version 2 1 6 and Skyline 2 5 When I run the QC with my data six replicate two conditions I get the error and see below Thanks Xi Data Processing for analysis 3 Error in structure colnames structure runID j more less view request Mascot Error Tolerant reposted as a new request 5 responses scappadona 2014 06 10 12 58 Dear Brendan I have a question regarding DAT files created with Mascot Error Tolerant search When working with the MS1 full scan filtering wizard I usually get a dialog box with all the modifications found in my DAT files In my current project I am working with DAT files created with Error Tolerant search in Mascot The DAT files seem to be correct and contain all the desired modifications in Unimod format Unfortunately when I import these data I don t get any message to add guessed modifications Furthermore when I inspect the library in the explorer I get a message that asks me to add only Carboxymethyl M 58 Can you tell me if I am missing some steps Am I wrong if I think that Skyline should automatically recognize and ask me to import the Unimod modifications in my DAT files as in the MS1 tutorial Might there be a specific problem with error tolerant searches Thanks for your support Salvo If you want to inspect an ErrorTolerant DAT files I could send you one by WeTransfer or similar more less view request Error in peptide numbering 1 response Dyrlund 2014 06 10 07 15 It seems that Skyline numbers all first and last amino acid of target peptides one less than the actual position in the protein sequence For example if I import the following into Skyline v 2 5 0 6079 Test RMGPKR Then selecting the first two peptides gives me RMGPK R 0 4 missed 1 R MGPK R 1 4 I would instead expect that the last peptide should be listed as R MGPK R 2 5 Is this an error or is there a reason for the different numbering compared to e g Uniprot more less PeptideNumbering png view request cannot download skyline any tips purnima mungalachetty 2014 06 06 07 07 more less view request Error specified libraries do not support the spectrum count peptide ranking 3 responses edammer 2014 06 05 16 21 I am getting this error when loading any of multiple sky files built from DDA runs on a QE plus where the mzXML and percolator filtered sorcerer sequest input files load fine whether 2 or up to 40 and allow me to work with the data and save the sky file with no problem but when I go back to skyline later and load the sky file I consistently get this error I am looking at 2 hour runs and limited the peptides per protein to 4 which required a ranking setting to be picked Libraries are blib files The exact message is The file contains an error on line 12 at column 8 Specified libraries do not support the Spectrum count peptide ranking more less view request Export of chromatogram images in a report 4 responses phalvey 2014 06 05 08 00 Hi guys Quick question We are trying to see if there is a way to export chromatogram images from multiple files as part of a report We have some folks who are working in a more regulatory enviroment and it s a big deal for them to have chromatograms which show where the peak was integrated and to include these images in a report along with peak areas etc I know individual images can be exported by right clicking and saving as an image file but is there any way to automate this and integrate them into a report Thanks and great work as always on the software Patrick Halvey more less view request Feature requests 4 responses Wouter Elings 2014 06 03 04 34 Dear Skyline team As I was trying to optimise my DIA methods I came up with some ideas that don t seem to be supported by Skyline These ideas are no use to anyone without a means to analyse the data so I hope you may be able to include support for it in a future release Alternatively it may be that Skyline is already able to handle my data but I haven t found the right settings I use Skyline 64 bit 2 5 0 5675 1 From your poster https skyline gs washington edu labkey files home software Skyline 2013 ASMS Overlapped DIA pdf I used the idea of 50 shifted MS2 frames in consecutive cycles From here it seems like a small step towards using for example 25 overlapping frames including four cycles in the isolation list before returning to the start or any other number other than one or two for that matter Yet Skyline does not seem to support this Upon importing the results I use Use results data isolation targets Deconvolution Overlap With these settings it can import the 50 version I assume the deconvolution works this way but when I try a 25 shifted method I get an error message Overlap deconvolution scheme is rank deficient at scan 136 Rank is 6 while matrix has dimension 7 A non degenerate overlapping window scheme is required Is it simply that this function is not supported in Skyline or am I missing something Relating to this I came up with a work around I figured I might be able to fool Skyline into thinking each 20mz MS2 window was actually an MSX 4 scan of four adjacent 5mz windows and let it do MSX deconvolution instead of overlap deconvolution Sadly it seems I can t import an isolation list and pasting windows in the Prespecified isolation windows field automatically orders the windows in increasing m z Would it be possible to have an isolation list import I think it would greatly enhance the user s possibilities of playing around with settings like these 2 If you take a DDA scan and plot hits m z values against elution time you ll notice a gradual shift in the distribution The hydrophobic peptides which elute later tend to have higher average precursor m z values Now when I m designing a DIA experiment I want its scan window to include as much data as possible while keeping the cycle time low The efficiency of doing this would increase if I were able to gradually shift my total isolation window over elution time For example scanning 220 720 mz at the beginning of the run and gradually shifting to 420 920 at the end I have made such a scan on a Q Exactive combining the 50 shifted cycles method with a stepwise increase over elution time using steps of my window size This step size ensures constant window placement which I think should enable demultiplexing However when I try to do this in Skyline upon importing I get the error message Failed to build a cache for file Tried to get a window mask for 730 a spectrum with previously unobserved isolation windows If possible I think it would be great if Skyline would be so versatile as to allow such method tweaks allowing for ever more efficient scanning Although I m no expert on coding I imagine that especially this second suggestion may be a rather tricky one Will you consider it Cheers Wouter more less view request Selecting and exporting optimal CE DP 1 response sstoychev 2014 06 02 03 52 I would like to find out if there is a way to select and more importantly export optimal CE DP values from a set of runs where CE DP were optimized i e samples analyzed with set of transition each with a range of CE DP values predetermined by Skyline Also is it possible to annotate MRM chromatographs showing the range of CE DP values 11 each with the actual values rather than 1 to 5 and 1 to 5 more less view request Background Proteome is there a maximum of Proteins 5 responses jrumsey 2014 05 28 16 29 I added a background proteome which consists of 195 650 proteins It seems to be taking quite a while to do the trypsin digest of the background proteome At the bottom of the screen it says resolving protein details for background proteome How do I know when it is done the green bar is only 1 2 way done When I look at unique peptides it seems to show the peptides but not showing any other proteins that share any of the peptides makes me wonder if it is not finished Any helpful hints more less view request Cannot export Scheduled isolation list anavare 2014 05 28 13 48 I am trying to use Skyline version 2 5 to set up a MRM method to run on a QE instrument To set up a test method I am using a simple BSA digest sample for which I have 2 DDA runs collected on QE I am having couple of issues to obtain a Scheduled isolation list from Skyline and was wondering if you could help in the troubleshoot Here is what I did the following steps 1 I first built a MS MS spectral library for the BSA digest using the two DDA runs interact pep xml files corresponding to the DDA files 2 Added the fasta sequence of BSA as a background proteome under Peptide setting Digestion 3 Then imported the fasta sequence of BSA into Skyline As expected the BSA peptides with corresponding MS MS data from the spectral library showed the MS2 spectra with highlighted fragment ions 4 Then I went to the Transition setting Full scan MS1 filtering and selected Count and Orbitrap and none or Targeted and Orbitrap for MS MS filtering 5 Next I exported the list as isolation list for Thermo Q extactive Here when I tried selecting Scheduled method I see the following error To export a scheduled method you must first choose the retention time predictor in peptide setting prediction or import results for all peptides in the document 6 I exported the list as Transition list with Standard method option The resulting csv file showed RT time for each peptide precursor 7 Then I went back to the Peptide setting Prediction and clicked Add Then I pasted the non redundant list of peptide precursors and RT obtained from the transition list csv file into the measured peptide window and clicked on Calculate The calculator did not auto calculate the slope and the y intercept as it did in one of the tutorials In fact I could not check the Auto calculate regression box since it is grayed out Thus I cannot predict the RT for the precursors and hence cannot generate a Scheduled isolation list to be exported on a QE instrument When I used the Standard method to export the isolation list I get blank columns for Start and End time even thought the RT time information is present in my spectral library Is there a way to fix this problem Am I doing something wrong more less view request Problem importing DIA raw file to skyline 1 response swee 2014 05 27 22 02 Hi I have used skyline to create a peptide spectral library from MaxQuant output of DDA runs of IEF fractions on QE The library has 5770 proteins 28500 peptides and 759630 transitions I have also carried out a multiplex MSX DIA run 5x4m z using QE on the unfractionated sample according to the method described in Nature Methods 2013 Vol 10 8 744 746 However I failed to import my DIA raw file as result into my skyline peptide spectral library Skyline hangs upon the import of my DIA raw file The transition settings that I have is exactly the same as the transition settings that I used for the generation of the isolation list for the multiplex 5x4m z DIA runs using skyline according to skyline mini tutorial that can be found at https brendanx uw1 gs washington edu labkey webdav home software Skyline 40files tips SkylineDIAMiniTutorial 2 1 pdf I m running skyline on Windows 7 Professional which has a RAM of 12 GB and it is a 64 bit operating system I m using Skyline 64 bit version 2 5 0 6079 Please help Thank you more less view request Add precursor peaks for targeted MS MS spectra PRM 2 responses Gerard 2014 05 27 06 09 Hi everybody This is probably a basic question but I don t see how to properly do what I m trying to do as I am doing some workaround I have data from a PRM experiment Q Exactive which therefore only has MS MS data Though I do see precursor peaks in those cases where the precursor has not been totally fragmented Then I know that I have several masses for the precursor that have been detected but it seems that I cannot add them in Skyline What I want I want to see those precursor chromatograms What I do and does not work the way I want I have the Full Scan configured for PRM this is None for MS1 filtering and Targeted Orbitrap 17 5K for MS MS filtering correct me if I am wrong somewhere I select the transitions for the peptide and I see one single mass of my precursor which I select I import the PRM raw file and I do see the precursor peak but I want to see all the forms of the precursor What I do to add all the precursors and kind of works I change the Full Scan configuration to Count Orbitrap 3 17 5K for MS1 filtering and the same as before for MS MS filtering I select the transitions for the peptide which now show all the variations for the precursor M 1 M M 1 2 etc Then go back to the Full Scan configuration and put again to None the MS1 filtering otherwise there is no precursor peaks extracted which makes perfectly sense as there is no MS1 information in the raw file Import then the file as before and I see all the precursors To re import a file I always have to remove the file SAVE and then import again Without saving or just marking for re import does make the job and just recovers the I suppose cached file I this a bug Sadly if you close and open again the Skyline all the precursors appear with the same mass and you have to repeat the operation for every peptide Then is there a better way to do what I am doing Many thanks in advance and for Skyline in general fantastic tool Gerard EDIT I also add that if you select the precursor with modifications like ammonia water losses they appear in the MS MS graph when you select the Split Graph section Not really related with the topic but still somehow similar more less view request Error The document format version 1 6 is not supported 1 response bouchal 2014 05 26 08 20 Hello I am getting crazy messages from Skyline in last few days Attempts to open some not all sky files or importing wiff files all since Friday result in the following error message The file contains an error on line 2 at column 2 The document format version 1 6 is not supported I am now using Skyline 2 5 0 5675 and Skyline daily 2 0 9 4899 at the same laptop Win 7 64 bit SP1 Where could be a problem Thanks Pavel more less view request the following modification could not be interpreted K 340 3 responses lorenzo vega 2014 05 23 15 38 Dear Brendan I have dealing with this issue for a few days without being able to find a solution I have created a library based on an ssl file attached There are a large number of peptides that have a K 340 modification Glucosylgalactosyl The modification is also added on the Peptide Settings menu When I build the library everything goes smooth but by the end Skyline send a mesage saying the following modification could not be interpreted K 340 The peptides with this modification are on the library but they do not have a peptide symbol on the side I created an small Skyline file where a peptide with this mod was added results file thermo raw where this peptide was identified was imported but I cannot get an ID mark on any of the XIC Could you indicate what is going on I am attaching some of and how can it be solved Best regards Lorenzo PS I have attached relevant files more less SpecialModsRegSearchLibraryWithOutTagReconRebuild blib SpecialModsRegSearchLibraryWithOutTagReconRebuild slc SpecialModsRegSearchLibrary ssl Test sky Test sky view Test skyd view request Exporting Data from Skyline Files 1 response ucnvtsf 2014 05 23 05 07 I have been having some trouble exporting specific data after measuring dynamic MRM of a BSA digest I am trying to validate my methods to measure dynamic MRM and therefore analysed the BSA digest at different concentrations I have succeeded in exporting information about the peak heights areas and retention times in excel files However the values of each of these measurements is exactly the same despite of the different concentrations used When I look at the transitions in the skyline files I do observe an increase in the intensity of each transition with increasing concentration Therefore I believe I might be doing something wrong during the export of the reports What might be the issue here more less view request Unique Peptide Search not working 1 response brian flatley 2014 05 22 07 52 Dear Brendan I was wondering if you could please help me with the Unique Peptides Search option as it is not working on the Skyline on my laptop however it is working on my colleagues laptop I am wondering if there is a setting or click on a box where I have to tick to get that working I am attaching two pictures herein first is showing how the search option looks on my lab mates skyline and the second image shows how it looks on my laptop s skyline I will be very grateful for your support Will be looking forward for your response Many Thanks Kind Regards more less unique peptides docx view request Is this normalization setting right 1 response yueg 2014 05 19 16 51 Dear Breadan Our data is from PSRM and I want to normalize the data phosphopeptide with non phosphopeptide when I right click the nonphosphopeptide and select the stdandard type as normalization I am wondering whether the software will use the total peak area of non phosphopeptide to normalize all other targeted phosphopeptide including each transition peak area or total peak transitions Also If I set more than 3 4 non phosphopeptide standard type as normalization The software will add all 3 4 non phosphopeptide peak area to normalize all other non phosphopeptide Is my understanding right Thank you for your kind help Eileen more less view request Modifications in specral library 3 responses hxli 2014 05 16 21 14 Hi I have build a spectra library directly from Maxquant search output msms txt with

    Original URL path: https://skyline.gs.washington.edu/labkey/project/home/support/begin.view (2014-08-11)
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  • Video 1: Method Editing: /home/software/Skyline
    using a browser with JavaScript disabled please enable it now Otherwise please update your version of the free Flash Player by downloading here Download the full video for faster access to repeat viewing recorded using Skyline v0 2 discussion Discussion Search Search Pages Skyline Targeted Proteomics Environment Install 32 bit Skyline v2 5 Install 32 bit Skyline v2 5 Unplugged Install 64 bit Skyline v2 5 Install 64 bit Skyline v2 5 Unplugged Release Notes Videos Video 1 Method Editing Video 2 Results Analysis Video 3 Existing Quantitative Experiments Video Trailer Tutorials 教程 中国语文 靶向 方法编辑 靶向方法优化 处理已有定量实验数据 MS1 全扫描筛选 靶向 MS MS PRM iRT 保留时间预测 チュトリアル 日本語版 ターゲットメソッドの編集 ターゲットメソッドの最適化 既存データ処理および定量実験 MS1フルスキャンフィルタ ターゲットMS MS PRM iRT保持時間予測 Targeted Method Editing Targeted Method Refinement Existing and Quantitative Experiments Absolute Quantification MS1 Full Scan Filtering Targeted MS MS PRM Custom and Live Reports Advanced Peak Picking Models iRT Retention Time Prediction Collision Energy Optimization Spectral Library Explorer QuaSAR Quantitative Statistics SRM Course Tutorials Tips Terminology Cheat Sheet In Progress How Skyline Builds Spectral Libraries ID Annotations Missing with Mascot Search Results DIA Methods for Thermo Q Exactive How Skyline Calculates Peak Areas and Heights Support for Bruker TOF Instruments AB SCIEX Instrument Settings

    Original URL path: https://skyline.gs.washington.edu/labkey/wiki/home/software/Skyline/page.view?name=video_0-2 (2014-08-11)
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