Figure 1.

Experimental setup and confirmation of the molecular identity of the target compound. Three sections of the study tissue were placed on a slide. One served as control, one was sprayed with three differently labeled versions (L1, L2, and L3) of the target compound, each at a different concentration (C1–C3), and a third tissue section was spotted with a seven-level (C1′–C7′) dilution series of L4. Finally, a fourth label (L4) was applied at the same concentration as L2 (C2) on both adjacent tissue sections for normalization purposes. The tissue studied is a pig colon tissue of which a hematoxylin and eosin (H&E) staining is shown in (b, left) and the MALDI-MSI image of the target histone H4 peptide is shown in (b, right). The identification of the target peptide was performed on tissue using high-mass accuracy delivered by a Q-exactive Orbitrap mass spectrometer with a mass error of 0.75 ppm (c), by a comparison of the observed and the expected isotopic patterns (d), and MS/MS experiments (Supporting Information Figure 1).