– DC4 crosslinker, an 18Å mass spectrometry-cleavable crosslinking reagent, features two intrinsic positive charges facilitating the fragmentation of cross-linked peptides into their constituent peptides through collision-induced dissociation or in-source decay. Initial fragmentation leads to cleavage adjacent to the positive charges, enabling the identification of cross-linked peptides as pairs of ions with precise mass separation. Furthermore, the resultant intact peptide fragments can undergo further fragmentation, producing a sequence of b- and y-ions for peptide identification.