Fig. 3: Human–yeast proteome mix to assess quantitative performance.
a–i, From diluted bulk digests, 250 pg of two-proteome mixes consisting of 150 pg of HeLa + 100 pg of yeast, 200 pg of HeLa + 50 pg of yeast and 240 pg of HeLa + 10 pg of yeast were injected. Peptides were separated at a throughput of 50 SPD. Data were recorded in DIA mode using optimal, but not the same, settings for the Orbitrap Astral mass spectrometer and Orbitrap Exploris 480 mass spectrometer and analyzed using DirectDIA+ in Spectronaut 18 at a 1% FDR. Quantification was performed at the MS1 level. Dots within the Bland–Altman plots (bottom) represent proteins with given log2 average PG abundance and log2 fold change of abundance across both proteome mixes. Density plots (right) depict the distribution of measured log2 fold change (FC) values, and CV diagrams (top) show the local CV of 100 proteins quantified with a rolling window over the entire abundance range; n = 3 technical replicates. For the Orbitrap Astral mass spectrometer, all quantified proteins (a, d and g) or only those proteins that were commonly quantified using the Orbitrap Exploris 480 mass spectrometer (b, e and h) are shown. For the Orbitrap Exploris 480 mass spectrometer, all quantified proteins are shown (c, f and i).
