Table 1.

Summary of MS acquisition methods DDA, DIA, and MRM/PRM with respect to precursor selection, precision of detection and quantification, reproducibility and PTM analysis.

Meth od	Precur sor selecti on	Fragme nt- ation	Collision method	Detectio n	Precision of peptide quantificati on	Reproducibilit y of peptide identification	Analysis (software)	PTM localization	Validation of PTM
DDA	Single	Single parent mass of the compound	CID, HCD, ETD, CAD51	All fragments ions26	Low/Moderate for spectra counts26	Low: missing peptides across multiple runs26	Multiple, straightforward178	CID: neutral loss ETD: multistage activation HCD: fragment ions	Predominant method with ETD fragmentation 15,179
DIA	Multiple	All peptides in a given m/z window	Alternating low-energy CID and high-energy CID51	All fragments ions29	Accurate: similar to MRM but more vulnerable to variation caused by interference from other peptides29	Moderate: ~80% peptide overlap across multiple runs180	Complex and requires deconvolution180,181	CID: neutral loss	Complex182
MRM	Single	Single parent mass of the compound	HCD51	High: Selection and detection of single fragment ion 29	Precise183	High38	Multiple, straightforward 184	MS2 fragmentation analysis185	Rely on synthetic unlabeled and stable isotope-labeled peptides80
PRM	Single of multiple	Single parent mass of the compound	CID HCD	All fragment ions	High: Run-to run peptide identification at the upper 85%39	High186	Multiple, straightforward	multistage activation fragmentation187	Useful to study PTMs that are low in abundance188