Mass Spectrometers can be operated in different modes, including data-dependent acquisition (DDA) mode (MS2 triggered by the most intense MS1 spectra), data independent acquisition (DIA) mode (windows of MS1 spectra are captured for MS2 analysis), and targeted mode (peptides are targeted based on a list of interest).

The two major Mass Spectrometry approaches to study the proteome are the top-down approach, a technique that analyzes intact proteins and enables near 100% sequence coverage but is challenging to scale, and the bottom-up approach, a technique for studying digested proteins before Mass Spectrometry analysis. While easy to implement, peptides must be computationally reconstructed into proteins for protein identification, and it has lower protein coverage than the top-down approach, making it more challenging to resolve proteoforms.