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Users’ manual

Main window
  1. Manage and compare searches: compare or delete existing data.
  2. Uploads: upload new datasets or databases.
  3. Result Basket: look at data you added to your result basket.

Manage and compare searches

Select searches

Select searches

Status can be “ok”, “error” or “loading”, meaning your results were loaded succesfully, your results were loaded with and error (click to see the error) or your results are being loaded. Select the checkboxes of the SearchIds you want to compare. Click on the Compare button to see all the proteins identified in the selected searches.

Compare searches and select protein

Select proteins

The order of the searches can be re-arranged by simple drag-and-drop. This feature is useful to group together searches which share the same experimental condition.

You can filter the list of proteins by choosing a post translational modification (PTM) of interest. Only the proteins containing this PTM will be kept.

Select a protein to switch to the detail view.

Protein identification details

Protein coverage view

Protein coverages

The coverages of the selected proteins are shown as green bars. The thickness of the bars correspond to the spectral counts for each peptide.

You can get more information about the amino acid sequence by moving with the mouse pointer over the graph. With a mouse-click on the green bars you get the Peptide Spectrum Match (PSM) details for the selected position.

Post Translational Modification (PTM) counts

PSM counts

By selecting a PTM from the drop-down menu you can see the positions with modifications as blue circles (only for the experimental data). The size of the blue circles corresponds to the spectral count containing the selected modification.

On top of the graph you can see labels indicating the modified amino acids (one letter code) together with their position. By clicking on either the labels or the blue circles you get the Peptide Spectrum Match (PSM) details for the selected position.

PSM zoom

You can zoom into a region by seleting an area with your mouse (press left button, select a region and release the button). The zoom can be reset with a double-click of the left mouse button in any empty area.

Peptide Spectrum Match (PSM) details

PSM details

Every PSM is represented by a gray line. By moving your mouse over the PSM’s you can see a popup showing information about the match (scan number, retention time, charge, precursor mass, AA sequence together with modifications and scores).

On the PSM’s you can see the selected PTM’s represented by circles:

  • Red circles show the most probable PTM positions. By “most probable” we mean the PTM positions found in the highest ranking PSM.
  • Orange circles indicate a conflicting position. This means the position was found in a lower ranking PSM, but has the same identification score as the highest ranking one.
  • Grey circles show PTM positions of lower ranking PSM’s.
  • Other modifications then the selected one are shown as black bars.

By clicking on any PSM you open a detail view showing the corresponding fragmentation spectrum and XIC’s.

Visualization of spectra and XIC’s

spectra and XIC details

By clicking on an PSM (gray lines) you open a view to the corresponding annotated fragmentation spectrum and XIC’s. This allows to easily verify the quality of the match and quantify the corresponding MS1 signals.

  1. Annotated fragmentation spectrum: the annotated fragmentation spectrum helps you to verify the quality of the match.
  2. Extracted Ion Chromatogram (XIC): quantify the corresponding MS1 signal.
  3. Alternative matches: look at alternative matches for this spectrum.

Annotated fragmentation spectrum

spectrum details

The spectrum shows annotated peaks with B-ions in blue and Y-ions in red. For phosphorylated PSM’s the fragments corresponding to neutral losses (98, 196, etc.) are shown in orange. You can zoom into a region by selecting an area with your mouse (left button).

To compare all opened fragmentation spectra amongst each other, you can click on the magnet button. This sets the mass range of all other spectra to the one from which you clicked. The zoom can be reset by clicking on the reload button by double-clicking on any empty area of the graph.

The zoom button opens the corresponding spectrum and XIC in an extra browser tab. You get a bigger view of both graphs. That is useful for looking at details and to make prettier screenshots.

Extracted Ion Chromatogram (XIC)

xic

The XIC is generated by extracting the MS1 signal of the given mass over charge (m/z) ratio and a tolerance of 10 ppm. Bars of different colors indicate MS2 scans found at the corresponding retention times:

  • Red: the MS2 event of the selected PSM.
  • Green: identified MS2 events with same precursor m/z and charge as the selected MS2.
  • Violet: identified MS2 events with same precursor mass and different.
  • Grey: non-identified MS2 events with same precursor mass.

Again, to compare all XIC’s amongst each other, you can click on the magnet button. This sets the retention-time range of all other XIC’s to the one from which you clicked. The zoom can be reset by clicking on the reload button or by double-clicking on any empty area of the graph.

Spectra and XIC’s can be closed by clicking on the close button.

xic quantitation

You can quantitate the XIC peaks by selecting a region. To do so you select an area, like done for zooming, but in this case you hold down the SHIFT key while selecting. A green area while show the region were the quantitation was done and a table containing intensity values will appear below.

The quantitation table shows the maximum intensity for the region of interest (Intensity) and the exact retention time where this intensity was found (Rt). This information can be added to the Result Basket by clicking on the basket button.

Uploads

Warning

The upload and delete functions are disabled for the public MsViz.

Upload results

Upload results

The types of data currently accepted by MsViz are either Mascot or MaxQuant results.

Data have to be uploaded as .zip files containing one or several results as described later in this section.

A MS1 intensity (minimum intensity value for precursor signal) can be set to increase upload speed.

By clicking the Upload button your .zip file will be loaded to MsViz. After a couple of seconds the new results should appear in Manage and compare searches. The parsing of the results and spectra information will take some time. You can see its status by pressing the refresh button in Manage and compare searches. Once the upload is finished, all Searches will be labelled with a green ok.

extract mzML from raw files

MsViz imports spectra data in the mzML format. You can extract mzML files from vendor specific raw files using MsConvert from ProteoWizard. Please make sure to activate the Peak Picking when extracting your raw files.

MsConvert

Mascot data

MsViz can read the mzIdentML results created by Mascot. You can export your Mascot results as mzIdentML (with file extension .mzid) directly from the interface using the default parameters.

You can upload .zip files containing one or several Mascot results together with their corresponding spectra in the .mzML format. There are 2 possibilities to group results together:

  1. usually the .mzid files from Mascot contain the name of the original raw spectra files. In this case MsViz will look for the same names, with the file extension .mzML (e.g. F004356.mzid together with the original spectra file 20150318_Sample_03.mzML).
  2. for resubmitted Mascot jobs the original raw spectra file names are missing. In this case the corresponding .mzML files have to have the same name as the .mzid files, with the .mzML file extension (e.g. F004357.mzid with F004357.mzML).

MaxQuant data

You can upload MaxQuant results as a .zip. file containing the following data:

  • the /txt folder from the MaxQuant results. You can directly use the whole folder, but MsViz will only parse information from the following tables, so you can remove the remaining tables if you like:

    • evidence.txt
    • msms.txt
    • parameters.txt
    • peptides.txt
    • proteinGroups.txt
    • summary.txt
  • the raw files mentioned in the summary.txt in the mzML format.

Warning

Only entries of type MULTI-MSMS in evidence.txt are imported to MsViz.

Upload database

Upload database

The database accepted by MsViz has to be in the FASTA format.

You can set special FASTA header parsing rules. By default MsViz will try to find the optimal parsing rules. But in case you want a special parsing or the parsing done by MsViz does not work, you can select one of the predefined rules and further refine them. There are the following predefined parsing rules available:

  • SwissProt_ID: parses the ID from SwissProt FASTA files
  • SwissProt_AC: parses the AC from SwissProt FASTA files
  • Trembl: parses Trembl FASTA files
  • custom: a custom setting where the first word after the > is selected

By clicking the Upload button your file will be loaded on MsViz.

You can see a list of the databases you already loaded.

Result Basket

Results basket

By clicking on any of the results in the list you open a view to the selected one.

You can remove your results or you can export them into a .csv containing the protein AC, the peptide, scan number, retention time, m/z, charge, score, localization score and intensity for all your samples.