Batch Processing

If you want to run READ for multiple input files sequentially, please install the requirements and follow the steps below.

TIP

This section assumes that you have read the previous sections on how to use READ! Minimal python knowledge is also beneficial!

Requirements

• Please install OpenMS.
  • [Recommended] We tested READ using OpenMS version 3.4.0!
• Please install uv.

Chimerys: Steps

• Put all your .raw files and identification files (e.g. PSMs and optionally proteins) in the same folder.
  • For example, let's call this folder tmt_files.
• PSM files should end in the suffix _PSMs.txt and be in tab-separated format.
• Protein files should end in the suffix _Proteins.txt and be in tab-separated format.
• For example, like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1_PSMs.txt
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1_Proteins.txt
• It's important that files that belong together are named like this schema.
• You will need to convert your .raw files to .mzML format, here's how to do it:
  • Download ThermoRawFileParser from here.
  • Convert your RAW files with:

    ThermoRawFileParser.exe -d path/to/tmt_files

• Running the multi-file scripts also requires that the resolution.csv from the Resolution GUI tool is in the same folder.
• Additionally, your config.toml should also be in the same folder. Adapt the configuration to your needs.
• Moreover, please put the tmt18plex_default.ini also in that folder and adapt if needed (see here).
• If you have a file with DIA mass windows, you should also put it in that folder.
  • You also have to set the filename in the tmt_chimerys_multi.py script under WINDOW_FILE.
  • You can ignore this for DDA results.
• Lastly, please put the following scripts into the folder:
  • multi/tmt_chimerys_multi.py
  • multi/tmt_chimerys_dda_multi.py
• Your tmt_files folder structure should now be something like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.mzML
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1_PSMs.txt
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1_Proteins.txt
  • ...
  • resolution.csv
  • config.toml
  • tmt18plex_default.ini
  • Mass List Table.csv (optional)
  • tmt_chimerys_multi.py
  • tmt_chimerys_dda_multi.py
• Open a terminal in this folder.
• Install uv if it's not already installed on your system, e.g.:

  pip install uv

• Run the DIA script with:

  uv run tmt_chimerys_multi.py

• or run the DDA script with:

  uv run tmt_chimerys_dda_multi.py

IMPORTANT

Please note that the same config file is used for all the files in the folder. The scripts automatically parse the window size from the MS file name and overrides the window size in the config file. You might have to adjust this parsing procedure if your filenames do not follow the same naming pattern. If a WINDOW_FILE is given, it will always use the windows from the file!

Spectronaut: Steps

• Put all your .raw files and identification file (e.g. precursors/the main report) in the same folder.
  • For example, let's call this folder tmt_files.
• The report file should end in the suffix .csv and be in comma-separated (.csv) format.
• For example, like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.csv
• Or like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • report.csv
• You need to set the filename of the identification file in the tmt_spectronaut_multi.py file, e.g. MAIN_REPORT="report.csv".
• You will need to convert your .raw files to .mzML format, here's how to do it:
  • Download ThermoRawFileParser from here.
  • Convert your RAW files with:

    ThermoRawFileParser.exe -d path/to/tmt_files

• Running the multi-file scripts also requires that the resolution.csv from the Resolution GUI tool is in the same folder.
• Additionally, your config.toml should also be in the same folder. Adapt the configuration to your needs.
• Moreover, please put the tmt18plex_default.ini also in that folder and adapt if needed (see here).
• If you have a file with DIA mass windows, you should also put it in that folder.
  • You also have to set the filename in the tmt_chimerys_multi.py script under WINDOW_FILE.
  • You can ignore this for DDA results.
• Lastly, please put the following scripts into the folder:
  • multi/tmt_spectronaut_multi.py
• Your tmt_files folder structure should now be something like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.mzML
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1_precursor.csv
  • ...
  • resolution.csv
  • config.toml
  • tmt18plex_default.ini
  • Mass List Table.csv (optional)
  • tmt_spectronaut_multi.py
• Open a terminal in this folder.
• Install uv if it's not already installed on your system, e.g.:

  pip install uv

• Run the Spectronaut script with:

  uv run tmt_spectronaut_multi.py

IMPORTANT

Please note that the same config file is used for all the files in the folder. The scripts automatically parse the window size from the MS file name and overrides the window size in the config file. You might have to adjust this parsing procedure if your filenames do not follow the same naming pattern. If a WINDOW_FILE is given, it will always use the windows from the file!

DIA-NN: Steps

• Put all your .raw files and the identification file (e.g. precursors/the main report) in the same folder.
  • For example, let's call this folder tmt_files.
• The report file should end in the suffix .parquet and be in parquet format.
• For example, like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.parquet
• Or like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • report.parquet
• You need to set the filename of the identification file in the tmt_diann_multi.py file, e.g. MAIN_REPORT="report.parquet".
• You will need to convert your .raw files to .mzML format, here's how to do it:
  • Download ThermoRawFileParser from here.
  • Convert your RAW files with:

    ThermoRawFileParser.exe -d path/to/tmt_files

• Running the multi-file scripts also requires that the resolution.csv from the Resolution GUI tool is in the same folder.
• Additionally, your config.toml should also be in the same folder. Adapt the configuration to your needs.
• Moreover, please put the tmt18plex_default.ini also in that folder and adapt if needed (see here).
• If you have a file with DIA mass windows, you should also put it in that folder.
  • You also have to set the filename in the tmt_chimerys_multi.py script under WINDOW_FILE.
  • You can ignore this for DDA results.
• Lastly, please put the following scripts into the folder:
  • multi/tmt_diann_multi.py
• Your tmt_files folder structure should now be something like this:
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.raw
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1.mzML
  • 20250519_Astral1_Evo_TH070_TT_THIDmulti003_pool_DIA_mz0c5_3ng_1_precursor.parquet
  • ...
  • resolution.csv
  • config.toml
  • tmt18plex_default.ini
  • Mass List Table.csv (optional)
  • tmt_diann_multi.py
• Open a terminal in this folder.
• Install uv if it's not already installed on your system, e.g.:

  pip install uv

• Run the DIA-NN script with:

  uv run tmt_diann_multi.py

IMPORTANT

Please note that the same config file is used for all the files in the folder. The scripts automatically parse the window size from the MS file name and overrides the window size in the config file. You might have to adjust this parsing procedure if your filenames do not follow the same naming pattern. If a WINDOW_FILE is given, it will always use the windows from the file!