Table 1.

Overview of features supported by different DIA software tools

Feature	SpectropectroNaut 18	DIA-NN 1.8.1	Open SWATH (part of OpenMS 3.0.0)	MSFragger-DIA 21	Skyline 23.1	EncyclopeDIA 2.12	MaxDIA 2.4
Free for Academia	✗	✓	✓	✓	✓	✓	✓
Open source	✗	✗	✓	✗	✓	✓	✗
Windows	✓	✓	✓	✓	✓	✓	✓
Linux	✓	✓	✓	✓	✓	✓	✓
Orbitrap	✓	✓	✓	✓	✓	✓	✓
TOFa	✓	✓	✓	✓	✓	✓	✓
DIA-PASEF	✓	✓	✓	✓	✓	✗	✓
PTM searches	✓	✓	✓	✓	✓	✓	✓
PTM localization scoring	✓	✓	✓	✓	✗	✓	✓
Multiplex (e.g., SILAC, mTRAQ)	✓	✓	✓	✓	✓	✓	✓
Interference removalb	✓	✓	✗	✓	✓	✓	✗
Visualization of MS data	✓	✗	✓	✓	✓	✓	✓
QC metric reporting	✓	✓	✓	✓	✓	✓	✓
Statistical analysis (diff. abundance)c	✓	✗	✗	✓	✓	✗	✗
No a priori knowledge DIA search (FASTA > DIA search)	✓	✓	✗	✓	✓	✓	✗
Library generation/refinement directly from DIA	✓	✓	✗	✓	✓	✓	✓
Library generation from DDA	✓	✗	✓	✓	✓	✓	✓
Combined library generation: DDA, DIA, GPF	✓	✗	✗	✓	✓	✓	✓
Identification/peak detection with neural networks/machine learning	✓	✓	✓	✓	✓	✓	✓
Integration of upstream tools for library generation	✓	✓	✓	✓	✓	✓	✓
Visualization/analysis capabilities of upstream tools’ search results	✗	✗	✗	✗	✓	✗	✗
Interactive visualization of data (personal opinion of the authors)	+ +		+	+	+ + +	+	+

DDA, data-dependent acquisition; DIA, data-independent acquisition; FAIMS, high-field asymmetric waveform ion mobility spectrometry; GPF, gas-phase fractionation; MS, mass spectrometry; PASEF, parallel accumulation and serial fragmentation; PTM, posttranslational modification; QC, quality control; SWATH, sequential window acquisition of all theoretical fragment ion spectra.

^aDepending on the specific software version and instrument file format combination, not all combinations might be possible for all software tools and MS vendors.

^bOpenSWATH and MaxDIA offer background subtraction.

^cBuilt-in functionality for differential statistics, to compare protein abundances between different samples/conditions.