SDRF-Proteomics: Data-independent acquisition (DIA) Template

1. Status of this Template

This document provides a template for annotating Data-independent acquisition (DIA) mass spectrometry experiments in SDRF-Proteomics format. This template defines additional columns that can be combined with any core template (human, vertebrates, etc.) to capture DIA-specific metadata.

Type: Experiment-type Template (used in combination with core templates)

Status: Released

Version: 1.1.0 - 2026-01

2. Abstract

Data-independent acquisition (DIA) is a mass spectrometry acquisition strategy where all precursor ions within a specified m/z range are fragmented systematically, regardless of their intensity. Unlike Data-dependent acquisition (DDA), DIA provides comprehensive and reproducible proteome coverage, reduced missing values across samples, and better quantitative accuracy. Popular DIA methods include SWATH-MS, MSE, All-Ion Fragmentation (AIF), and various windowed DIA approaches.

DIA experiments have unique characteristics: predefined m/z isolation windows, scan window ranges, stepped collision energies, and spectral library requirements. This template captures the acquisition parameters essential for DIA data analysis and reproducibility.

3. Connections to Analysis Pipelines

DIA data is typically analyzed using specialized software:

DIA-NN: Deep learning-based DIA analysis
Spectronaut: Commercial DIA analysis platform
OpenSWATH: Open-source targeted analysis
MaxDIA: MaxQuant’s DIA module
EncyclopeDIA: Chromatogram library search
Skyline: Targeted quantification

When documenting DIA experiments, noting the intended or used analysis software is RECOMMENDED.

4. Checklist

This section defines the metadata columns required and recommended for DIA experiments.

4.1. Recommended Columns

The following columns are RECOMMENDED for DIA experiments (in addition to the core SDRF-Proteomics requirements):

Column Requirement Description Ontology/CV Example Values

comment[scan window lower limit] RECOMMENDED Lower m/z limit of the DIA scan range Numeric value (m/z) 400, 350

comment[scan window upper limit] RECOMMENDED Upper m/z limit of the DIA scan range Numeric value (m/z) 1200, 1650

comment[isolation window width] RECOMMENDED Width of DIA isolation windows PSI-MS (MS:1000828) 25, 8, variable

comment[number of isolation windows] RECOMMENDED Total number of DIA windows per cycle Numeric value 32, 64, 100

comment[collision energy] RECOMMENDED Collision energy used for fragmentation (can have multiple for stepped CE) PSI-MS (MS:1000045) 27 NCE, 30 eV, 22 NCE;26 NCE;30 NCE

comment[dissociation method] RECOMMENDED Fragmentation method used PRIDE (MS:1000044) NT=HCD;AC=PRIDE:0000590

comment[MS2 mass analyzer] RECOMMENDED Mass analyzer used for MS2 acquisition PSI-MS (MS:1000443) NT=Orbitrap;AC=MS:1000484

comment[cycle time] RECOMMENDED Time to complete one DIA cycle Numeric value with unit 3.2 s, 2.8 s

comment[spectral library] RECOMMENDED Spectral library used for analysis Free text or accession project-specific library, Pan-Human library, PRIDE:PXD000001

comment[DIA method] RECOMMENDED Specific DIA method variant (child of PRIDE:0000450) PRIDE (PRIDE:0000450) NT=SWATH-MS;AC=PRIDE:0000451, diaPASEF, scanning SWATH

Column	Requirement	Description	Ontology/CV	Example Values
`comment[scan window lower limit]`	RECOMMENDED	Lower m/z limit of the DIA scan range	Numeric value (m/z)	400, 350
`comment[scan window upper limit]`	RECOMMENDED	Upper m/z limit of the DIA scan range	Numeric value (m/z)	1200, 1650
`comment[isolation window width]`	RECOMMENDED	Width of DIA isolation windows	PSI-MS (MS:1000828)	25, 8, variable
`comment[number of isolation windows]`	RECOMMENDED	Total number of DIA windows per cycle	Numeric value	32, 64, 100
`comment[collision energy]`	RECOMMENDED	Collision energy used for fragmentation (can have multiple for stepped CE)	PSI-MS (MS:1000045)	27 NCE, 30 eV, 22 NCE;26 NCE;30 NCE
`comment[dissociation method]`	RECOMMENDED	Fragmentation method used	PRIDE (MS:1000044)	NT=HCD;AC=PRIDE:0000590
`comment[MS2 mass analyzer]`	RECOMMENDED	Mass analyzer used for MS2 acquisition	PSI-MS (MS:1000443)	NT=Orbitrap;AC=MS:1000484
`comment[cycle time]`	RECOMMENDED	Time to complete one DIA cycle	Numeric value with unit	3.2 s, 2.8 s
`comment[spectral library]`	RECOMMENDED	Spectral library used for analysis	Free text or accession	project-specific library, Pan-Human library, PRIDE:PXD000001
`comment[DIA method]`	RECOMMENDED	Specific DIA method variant (child of PRIDE:0000450)	PRIDE (PRIDE:0000450)	NT=SWATH-MS;AC=PRIDE:0000451, diaPASEF, scanning SWATH

Note	The parent term for all DIA methods is Data-independent acquisition (PRIDE:0000450). Use `comment[DIA method]` to specify the specific variant when known. Check the PRIDE Controlled Vocabulary for available terms.

4.2. Optional Columns

The following columns are OPTIONAL but commonly used:

Column Requirement Description Ontology/CV Example Values

comment[precursor mass tolerance] OPTIONAL Mass tolerance for precursor ions in database search Numeric value with unit 10 ppm, 20 ppm

comment[fragment mass tolerance] OPTIONAL Mass tolerance for fragment ions in database search Numeric value with unit 0.02 Da, 20 ppm

Column	Requirement	Description	Ontology/CV	Example Values
`comment[precursor mass tolerance]`	OPTIONAL	Mass tolerance for precursor ions in database search	Numeric value with unit	10 ppm, 20 ppm
`comment[fragment mass tolerance]`	OPTIONAL	Mass tolerance for fragment ions in database search	Numeric value with unit	0.02 Da, 20 ppm

5. Stepped Collision Energy

Many DIA methods use stepped or ramped collision energies to improve fragmentation coverage. Document this using multiple comment[collision energy] columns or semicolon-separated values:

5.1. Multiple Columns Approach

comment[collision energy]	comment[collision energy]	comment[collision energy]
22 NCE	26 NCE	30 NCE

comment[collision energy]

22 NCE

26 NCE

30 NCE

5.2. Semicolon-Separated Approach

comment[collision energy]
22 NCE;26 NCE;30 NCE

comment[collision energy]

22 NCE;26 NCE;30 NCE

Both approaches are valid. Use the approach that best fits your workflow.

6. Isolation Window Schemes

DIA methods vary in their isolation window strategies:

Scheme	Description	How to Document
Fixed windows	Equal-width windows across mass range	comment[isolation window width] = 25
Variable windows	Windows optimized for ion density	comment[isolation window width] = variable; provide method file reference
Overlapping windows	Windows with m/z overlap	Note in comment[DIA method] or separate documentation
Scanning/sliding windows	Continuous scanning approach	comment[DIA method] = scanning SWATH

Scheme

Description

How to Document

Fixed windows

Equal-width windows across mass range

comment[isolation window width] = 25

Variable windows

Windows optimized for ion density

comment[isolation window width] = variable; provide method file reference

Overlapping windows

Windows with m/z overlap

Note in comment[DIA method] or separate documentation

Scanning/sliding windows

Continuous scanning approach

comment[DIA method] = scanning SWATH

For complex schemes, reference the instrument method file in comment[method file] if available.

7. DIA Method Variants

Common DIA acquisition methods and their characteristics:

Method	PRIDE CV	Vendor/Origin	Key Features	Example Annotation
SWATH-MS	PRIDE:0000451	SCIEX	Fixed or variable windows, typically 25-100 windows	NT=SWATH-MS;AC=PRIDE:0000451
MSE	PRIDE:0000452	Waters	All-ion fragmentation, alternating low/high energy	NT=MSE;AC=PRIDE:0000452
All ion fragmentation (AIF)	PRIDE:0000453	Thermo	All-ion fragmentation approach	NT=All ion fragmentation;AC=PRIDE:0000453
diaPASEF	(pending)	Bruker	Ion mobility-enhanced DIA	diaPASEF
boxcar DIA	(pending)	Various	Multiplexed acquisition with notched windows	boxcar DIA
Scanning SWATH	(pending)	SCIEX	Continuously scanning isolation window	scanning SWATH

Method

PRIDE CV

Vendor/Origin

Key Features

Example Annotation

SWATH-MS

PRIDE:0000451

SCIEX

Fixed or variable windows, typically 25-100 windows

NT=SWATH-MS;AC=PRIDE:0000451

MSE

PRIDE:0000452

Waters

All-ion fragmentation, alternating low/high energy

NT=MSE;AC=PRIDE:0000452

All ion fragmentation (AIF)

PRIDE:0000453

Thermo

All-ion fragmentation approach

NT=All ion fragmentation;AC=PRIDE:0000453

diaPASEF

(pending)

Bruker

Ion mobility-enhanced DIA

diaPASEF

boxcar DIA

(pending)

Various

Multiplexed acquisition with notched windows

boxcar DIA

Scanning SWATH

(pending)

SCIEX

Continuously scanning isolation window

scanning SWATH

Note	Methods marked as "(pending)" do not yet have PRIDE CV accessions. Use the free text name until official terms are available.

8. Example SDRF File

A complete example for a DIA proteomics experiment:

source name	characteristics[organism]	characteristics[disease]	...	comment[proteomics data acquisition method]	comment[scan window lower limit]	comment[scan window upper limit]	comment[DIA method]	comment[data file]
plasma_sample_001	homo sapiens	normal	...	Data-independent acquisition	350	1650	SWATH-MS	plasma_001.raw
plasma_sample_002	homo sapiens	normal	...	Data-independent acquisition	350	1650	SWATH-MS	plasma_002.raw
plasma_sample_003	homo sapiens	type 2 diabetes mellitus	...	Data-independent acquisition	350	1650	SWATH-MS	plasma_003.raw

Sample metadata Data file metadata

Note	The `…` column indicates omitted columns (organism part, biological replicate, technology type, label, instrument, cleavage agent, fraction identifier, technical replicate, isolation window width, collision energy, mass tolerances, dissociation method).

9. diaPASEF Example

For ion mobility-enhanced DIA (diaPASEF), additional columns may be relevant:

Column Name	Description	Example Values
comment[ion mobility]	Ion mobility separation used	TIMS, FAIMS, not applicable
comment[1/K0 range]	Ion mobility range	0.6-1.6 1/K0

Column Name

Description

Example Values

comment[ion mobility]

Ion mobility separation used

TIMS, FAIMS, not applicable

comment[1/K0 range]

Ion mobility range

0.6-1.6 1/K0

10. Best Practices for DIA Annotation

Always specify acquisition method: Use comment[proteomics data acquisition method] with the PRIDE CV term for DIA.
Document scan range: Include comment[scan window lower limit] and comment[scan window upper limit] for reproducibility.
Report isolation window details: Document window width and number of windows when known.
Include collision energy: Document all collision energies used, especially for stepped CE methods.
Note the DIA variant: Use comment[DIA method] to specify SWATH-MS, diaPASEF, MSE, etc.
Reference spectral libraries: If using a spectral library, document it in comment[spectral library].
Report mass tolerances: These are essential for data reanalysis.
Consider instrument-specific parameters: Different platforms may have unique parameters worth documenting.

11. Template File

The DIA SDRF template file is available in this directory:

dia-acquisition-template.sdrf.tsv

12. Validation

DIA SDRF files should be validated using the sdrf-pipelines tool:

pip install sdrf-pipelines
parse_sdrf validate-sdrf --sdrf_file your_file.sdrf.tsv

Note	DIA-specific validation rules are under development.

13. Authors and Maintainers

This template was developed by the SDRF-Proteomics community with contributions from DIA proteomics researchers.

For questions or suggestions, please open an issue on the GitHub repository.