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Messages - UFZ_Stef

17
MS-DIAL / Re: The use of RI for Alignment and Identification
Dear Hiroshi,

thanks for your giving insights into your routine! This is the way I do it as well. So there must be something in my alkane dictionary.

Now that you reported on the fact that MS Dial does not need the alkane in each sample I will investigate my problem more deeply.

Thank you for your assistance and the work I you are investing into the tool,

Stefan
20
MS-DIAL / The use of RI for Alignment and Identification
Dear appreaciated Forum,

Our instrument has a stable performance and the retention does not shift dramatically (~4 seconds in 250 runs). We had just a few samples which contained alkanes for setting up an alkane dictionary. Here the alkane mix was added to the sample after the first GC Injection to avoid co elution. Most of our samples do not contain alkanes. In conclusion we consider the RT as an effective alignment parameter which I want to use for the alignment (which I have discussed here: http://www.metabolomics-forum.com/index.php?topic=1570.0).

But:

MS Dial seems to expect the presence of alkanes from the dictionary in each sample, right? So far I haven't received reliable RI values for samples that do not contain the alkanes themselves. (I took the Retention Time of the alkane dictionary from an representative sample of the same sequence).



For identification purposes the RI is an important parameter. But do I really need alkanes in each sample if there was no severe shift in the retention time? In my oppinion libraries have greater variation in RI (due to differences chromatography) than samples from one sequence. So there is no need to have a very precise RI estimate for each peak when comparing this to a highly variable RI from the library.


I looking forward to your comments. Do you always add alkanes to all samples? Do you prepare an alkane dictionary for each sample individually (since there are always RT differences in the alkane peaks among two samples) or how does MS Dial detects the alkane peaks in each sample?

Kind Regards,
Stefan
21
MS-DIAL / Re: Huge Retention Time differences in alignment at narrow retention time tolerance
Even though replying to one's own comments is like a one man show I would like to further comment on the problem I have described above  :D

The high difference in the RT of the aligned peaks is due to wrongly determined RI values, which I actually not even worked with. To my mind there are several error's in MS Dial I would like to report on:


Run 1:

In my MS Dial Analysis parameter setting I avoided using RI for Peak identification and peak alignment. Please have a look to the screenshots 4 and 5. Nevertheless MS-Dial requests for a RI dictionary (which is not needed in this case) by the end of the parameter setting menu and without uploading a file I could not start the analysis. This is shown in screenshot 6. So I uploaded a dictionary (even though my samples do not contain alkanes) and the analysis started. Keep in mind that I still had 'Use retention time' checked!

Results: There were just a few peaks aligned and the time axis of the Alignment spot viewer was displayed in RI (which were wrongly calculated). So MS Dial ignored my 'Use retention time' setting and still tried to align on RI basis. The wrong RI calculation is the reason why there were huge time differences in RT (because the wrong RI matched well).

Conclusion: MS Dial automatically selects RI alignment over RT, even though the user made a different selection. By the way I am also wondering why MS-Dial does the Gap filling automatically, even though I haven't selected this option (see screenshot 5.PNG and Gap_filling_but_unchecked.PNG).

Run 2:

2) After the first alignment I had a second look to the Analysis parameter setting (in the same project) and I switched back to 'Use retention time'. Then I ran the analysis for a second time. Now the Alignment Spot Viewer has the RT on the x axis an the aligned peaks were much closer since the algorithm really used the RT setting I made.



I am looking forward for responses and I also saw developers of MS Dial around. Please have a look to the issue I have described here.

Kind regards,

Stefan
22
MS-DIAL / Huge Retention Time differences in alignment at narrow retention time tolerance
Hey Everyone,

I have discovered something weird in the MS Dial tool and I am highly interested in your experiences with following issue:

 I used the retention time for alignment and the retention time tolerance was set to 0.075 min (4.5 seconds). Then the exported retention time matrix of the aligned peaks was checked for differences in retention time (min, max, range) and I have discovered that there were Peaks aligned which were several minutes apart. Please have a look to the screenshot of the export RT table: high_diff_in_RT.PNG. In the frist line I marked a peak which has a RT of 7.9 min and this was aligned with 5.9 min RT peaks... This is even worse than the statistical compare tool of LECO and I am very sceptical that this is the usual output. Has anyone experienced the same? Why should  set up a tolerance value if this is ignored by the aligning algorithm?!

Please have a look to your data and tell me if you have experienced similar results!

Kind Regards
Stefan
23
MS-DIAL / MS Dial Problem in processing the demo dataset
Dear Metabolomics Community,

great to a format like this for asking questions. I just started playing around with MS DIAL and the tool looks very promising to me (at least the software includes all the steps I did with my data on a different platform).

But when I process the demo dataset "Alkanes Kovats deom by Arabidopsis" I fail to achieve the same results as it is shown in the tutorial. I am using Version 4.24 on a Win10 and I followed the tutorial carefully but Alignment Spot Viewer stays empty and also the Rep. vs Ref. Window does not show any Refence (but instead "No information"). It seems that the "GCMS DB_MassBank-Restek-RIKEN.msp" was not considered for comparison. May someone else try to run the Demo dataset? If I use RT instead of RI for the alignment I obtain results for the alignment but the Identification remains open. So the problem might be related to the usage of RI.

I also tried to align data that I have obtained myself and the Alignment Spot viewer gave reasonable results. Therefore the software itself seems to work fine...

Has anyone experienced similar problems?

Kind regards
Stefan