Hello Alex, Glad that you were able to get to the bottom of this issue and hope you have been able to process your data.
I just wanted to add that when installing TPP on Windows, the entire ProteoWizard set of tools (including msConvert GUI) is also bundled -- so there is typically no need to re-download/install unless you want a newer version. You can find the software under C:/Program Files/ProteoWizard/ on standard installs. Note that one is able to install multiple versions side-by-side, and you may even notice multiple ones there already if you have updated TPP on that machine before. Cheers, --Luis On Thu, Jan 18, 2024 at 7:53 AM Alex Gao <alexkao1...@gmail.com> wrote: > Dear David, > > So after a long troubleshooting process with msConvert and Waters team as > well as testing it by myself, I found the best way to convert Waters.raw > files is to use this Waters proprietary software > https://microapps.on-demand.waters.com/home/showmarkdown/data-as-a-product > to convert Waters file to mzML (convert to 32 bit). Alternatively, one > could convert the file with msConvert GUI (the actual software, not the one > embedded in TPP), with only the settings "write index" and "TPP compatible" > checked, convert to mzML 32-bit (mzXML 32-bit for Maxquant compatibility), > and most importantly, exclude the negative values (which are EMR spectra > from UV or PDA detector inline with the system) by adding a subset filter > for MS-Level "1-", which allows the mz(X)ML to keep only MS1 and MS2 data. > Thanks for your help! > > Best, > Alex > On Thursday, January 11, 2024 at 2:40:07 AM UTC+1 David Shteynberg wrote: > >> Hello Alex, >> >> This is pointing to a bug in either the Waters library for access to the >> raw data or in msconvert. Perhaps you can contact Waters and msconvert >> developers to see if they have patched the problem. My suspicion as to the >> nature of the problem is use of signed (as opposed to unsigned) variables >> internally for the representation of intensities. Once the intensities get >> large enough they roll over to the negative representations of the values >> in C++. Let us know when you find a solution to this issue! >> >> Thanks, >> -David >> >> On Jan 10, 2024, at 4:24 AM, Alex Gao <alexk...@gmail.com> wrote: >> >> Hi David, >> >> I do not know why. The data was acquired on MassLynx by a Waters Xevo >> QTOF set to fast DDA, with scan times of MS1 and MS2 set to both 1s with 10 >> MS2 scans. The lockspray mass is also included. The settings, besides the >> scan times, were not changed from the last time I ran the data, which >> worked the last time. On MassLynx, the intensities look normal as well, >> with a dome of increasing intensities over the gradient, typical of bottom >> up proteomics. Is there a way to solve this? >> >> On Wednesday, January 10, 2024 at 1:56:19 AM UTC+1 David Shteynberg wrote: >> >>> Hello Alex, >>> >>> I was able to download your file, convert the data to mzML using >>> msconvert and read a few spectra using the command line tool readMzXml >>> (works on mzML too). Unfortunately, I am finding many intensities these >>> files that are encoded with negative values, which is likely causing >>> problems for the downstream tools like comet. Do you know why negative >>> intensities are in these files? It is possible this is a bug with the raw >>> encoding. Are you able to visualize these files using Waters software to >>> check the intensities and compare to those output by msconvert? >>> >>> Thanks, >>> -David >>> >>> On Jan 5, 2024, at 4:17 AM, Alex Gao <alexk...@gmail.com> wrote: >>> >>> Hi David, >>> >>> Thank you for your prompt feedback! The file can be found at the link >>> below. >>> >>> >>> https://drive.google.com/file/d/1QulcwZ3UP7pcMtECNwubd4DysqaDq4UN/view?usp=sharing >>> >>> Looking forward to your inputs! >>> >>> Best, >>> Alex >>> >>> On Friday, January 5, 2024 at 1:09:03 AM UTC+1 David Shteynberg wrote: >>> >>>> Hello Alex, >>>> >>>> Thanks for trying the TPP pipeline to solve your proteomics >>>> computational needs. Sorry, you are having trouble converting and >>>> analyzing Waters raw data. Have you tried using the latest version of >>>> msconvert to do the conversion to mzML? If you are able to upload your >>>> file to an online drive and give me access to it, I can pull it down and >>>> try to convert it myself. Let me know what works for you. >>>> >>>> Cheers! >>>> -David >>>> >>>> On Jan 3, 2024, at 4:32 AM, Alex Gao <alexk...@gmail.com> wrote: >>>> >>>> Hi guys, >>>> >>>> So I've used TPP before to convert bottom up proteomic Thermo raw files >>>> in msconvert, comet search with custom params file, and xinteract to >>>> perform protein prophet with no issues. Earlier this year, I performed the >>>> same with Waters. Raw file, although in the beginning there was an issue >>>> with size, shrinking the size of my raw file from 25 to 5-6GB did the >>>> trick. >>>> >>>> However, recently, I reinstalled TPP (v6.3.3), and was trying to >>>> perform the same task to convert 5-6GB of Waters.raw but failed. msconvert >>>> didn't fail, but comet search was finished in less than 1min, which is not >>>> quite right. And out of the 200,000 spectra in the original file, only 1000 >>>> ish was used for comet search. this obviously leads to an error code of 256 >>>> at the xinteract level. >>>> >>>> I tried several different troubleshoots, not changing the default >>>> params file except for location of fasta file, uninstalling and >>>> reinstalling, changing a computer, renaming the FUNC0003 files from my >>>> Water.raw directory for the search, etc. etc., but none work. >>>> >>>> Any help would be much appreciated. Thank you! >>>> >>>> Best, >>>> Alex >>>> >>>> -- >>>> You received this message because you are subscribed to the Google >>>> Groups "spctools-discuss" group. >>>> To unsubscribe from this group and stop receiving emails from it, send >>>> an email to spctools-discu...@googlegroups.com. >>>> To view this discussion on the web visit >>>> https://groups.google.com/d/msgid/spctools-discuss/84cf1e94-32b3-47f6-80a8-37150ff94d93n%40googlegroups.com >>>> <https://groups.google.com/d/msgid/spctools-discuss/84cf1e94-32b3-47f6-80a8-37150ff94d93n%40googlegroups.com?utm_medium=email&utm_source=footer> >>>> . >>>> >>>> >>>> >>> -- >>> You received this message because you are subscribed to the Google >>> Groups "spctools-discuss" group. >>> To unsubscribe from this group and stop receiving emails from it, send >>> an email to spctools-discu...@googlegroups.com. >>> >>> To view this discussion on the web visit >>> https://groups.google.com/d/msgid/spctools-discuss/d774140c-2641-48f2-b9c7-900442b10503n%40googlegroups.com >>> <https://groups.google.com/d/msgid/spctools-discuss/d774140c-2641-48f2-b9c7-900442b10503n%40googlegroups.com?utm_medium=email&utm_source=footer> >>> . >>> >>> >>> >> -- >> You received this message because you are subscribed to the Google Groups >> "spctools-discuss" group. >> To unsubscribe from this group and stop receiving emails from it, send an >> email to spctools-discu...@googlegroups.com. >> >> To view this discussion on the web visit >> https://groups.google.com/d/msgid/spctools-discuss/6d911bb1-1bc9-4135-aafa-b168657ad219n%40googlegroups.com >> <https://groups.google.com/d/msgid/spctools-discuss/6d911bb1-1bc9-4135-aafa-b168657ad219n%40googlegroups.com?utm_medium=email&utm_source=footer> >> . >> >> >> -- > You received this message because you are subscribed to the Google Groups > "spctools-discuss" group. > To unsubscribe from this group and stop receiving emails from it, send an > email to spctools-discuss+unsubscr...@googlegroups.com. > To view this discussion on the web visit > https://groups.google.com/d/msgid/spctools-discuss/a4ed1d74-04a0-468d-b8a4-deae9bfc5e69n%40googlegroups.com > <https://groups.google.com/d/msgid/spctools-discuss/a4ed1d74-04a0-468d-b8a4-deae9bfc5e69n%40googlegroups.com?utm_medium=email&utm_source=footer> > . > -- You received this message because you are subscribed to the Google Groups "spctools-discuss" group. 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