Novel cross-links between an oxidized histidine and intact histidine, lysine, or cysteine residues were discovered and characterized from high-molecular weight (HMW) fractions of an IgG1 monoclonal antibody (mAb). The mAb HMW fractions were collected using preparative size-exclusion chromatography (SEC) and extensively characterized to understand the mechanism of formation of the nonreducible and covalently linked portion of the HMWs. The HMW fractions were IdeS digested, reduced, and analyzed by size-exclusion chromatography coupled with mass spectrometry (SEC-MS). The nonreducible cross-links were found to be enriched in the fragment crystallizable (Fc) region of the heavy chain, with a net mass increase of 14 Da. Detailed peptide mapping revealed as many as seven covalent cross-links in the HMW fractions, where oxidized histidines react with intact histidine, lysine, and free cysteine to form cross-links. It is the first time that histidine-cysteine (His-Cys) and histidine-lysine (His-Lys) in addition to histidine-histidine (His-His) cross-links were discovered in monoclonal antibody HMW species. The histidine oxidation hot spots were identified, which include conserved histidine residues His292 and His440 in the Fc region and His231 in the hinge region of the IgG1 mAb heavy chain. Their cross-linking partners include His231, His292, His440, and Cys233 in the hinge region and Lys297 in the Fc region. A cross-linking mechanism has been proposed that involves nucleophilic addition by histidine, cysteine, or lysine residues to the carbonyl-containing histidine oxidation intermediates to form the cross-links.