The CHIMERYS processing workflow for data independent acquisition does not require a quantitation node. It receives quantitative values directly from CHIMERYS for DIA and PRM data. The DIA processing workflow is shown in the following figure.

CHIMERYS DIA processing workflow
CHIMERYS DIA processing workflow

The CHIMERYS node has two new parameters that are important for data independent analysis. The parameters are as follows:

  • Processing Mode: This parameter controls the MBR feature of CHIMERYS.
    • One Job per File: If this option is selected, then MBR is not activated and the results only include IDs from each file run independently.
    • One Job for All Files: If this option is selected, then MBR is activated.

    • NOTE

      Due to memory constraints, the One Job for All Files option will fail with large datasets greater than 100 GB. Therefore, to perform MBR with large datasets, Thermo Fisher Scientific recommends using the CHIMERYS Inclusion File.

    • One Job per Fraction in File: If this option is selected, the application performs independent CHIMERYS runs for each file regardless of it belonging to fractionated data or not.
  • CHIMERYS Inclusion File: This parameter lets you select a CHIMERYS Inclusion File.
  • The CHIMERYS Inclusion File performs MBR one file at a time using the PSMs in the inclusion file as the targets to try to quantify. Thermo Fisher Scientific recommends using this parameter for very large datasets where the One Job Per File parameter is not a possible option.

The CHIMERYS Inclusion File can be exported from any CHIMERYS .msf file or .pdResult file using File > Export > To CHIMERYS Inclusion file. The CHIMERYS inclusion file must only include PSMs from the peptides you would like to quantify across all runs. Adding too many PSMs to the inclusion file might negatively affect the number of identified and quantified peptides.

The Consensus workflow includes a new node called the Fragment Ions Quantifier node. You must use this node to quantify DIA and PRM datasets. The Fragment Ions Quantifier node works similarly to the Precursor Ions Quantifier node and Reporter Ions Quantifier node.

The default DIA quantification workflow CWF_Comprehensive_Enhanced_Annotation_DIA is shown in the following figure.

CWF_Comprehensive_Enhanced_Annotation_DIA default workflow
CWF_Comprehensive_Enhanced_Annotation_DIA default workflow

There are no parameters in the Fragment Ions Quantifier node that are only specific for DIA or PRM analysis. For a detailed description of the node parameters, see Fragment Ions Quantifier node in the Proteome Discoverer Help.