Heralds of parallel MS: Data-independent acquisition surpassing sequential identification of data dependent acquisition in proteomics [Research]

Comprehensive, reproducible and precise analysis of large sample cohorts is one of the key objectives of quantitative proteomics. Here, we present an implementation of data-independent acquisition using its parallel acquisition nature that surpasses the serial limitation of data-dependent acquisition on a quadrupole ultra-high field Orbitrap mass spectrometer. In deep single shot data-independent acquisition, we quantified over 7,500 proteins of human cell lines and 9,000 proteins of mouse tissues, with fewer than 2.5% missing proteins and median coefficients of variation of down to 5% among replicates. In very complex mixtures, we could quantify more than 15,000 proteins, outperforming data-dependent acquisition methods by more than two-fold. Using this method, we investigated large-protein networks before and after the critical period for whisker experience-induced synaptic strength in the murine somatosensory cortex 1 barrel field. This work shows that parallel mass spectrometry enables proteome profiling for discovery with high coverage, reproducibility, precision and scalability.

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