A Potential Mercury Polymer Mass Tag Probe for Mass Cytometry

A novel mercury(II) polymer with pendant dipicolylamine (DPA) chelators carrying Hg²⁺ ions was prepared as a potential mass tag for mass cytometry (MC) applications. The polymer itself, with DP ≈ 22 had a mixture of PEG₂₄ (provides stealth) and PEG₂₃-azide groups attached to the pendant groups, with ca. 3 azide groups per polymer for antibody conjugation. The initial Hg-DPA complex had 2 Cl atoms attached to each Hg²⁺ ion (Cl₂-Hg-DPA). An approach to replace the two Hg-Cl bonds via ligand exchange reactions by treating the Hg-DPA polymer with monothiol ligands (glutathione, cysteine, and thioglycolic acid) led to a loss of Hg, while Hg polymers treated with dithiol ligands (2,3-dimercapto-1-propanol and 2,3-dimercapto-1-propanesulfonic acid sodium salt monohydrate) led to polymers <b>2d</b> and <b>2f</b>, respectively, in which both polymers largely retained their Hg content. These polymers showed relatively low levels of nonspecific binding to suspensions of peripheral blood mononuclear cells (PBMCs). Both polymers were conjugated to anti-CD8a via strain-promoted azide-alkyne coupling, and the conjugates were employed in a 10-plex assay of PBMCs along with Maxpar reagents. Polymer <b>2d-CD8a</b> showed excellent specificity to target CD8a antigens, even at a low titer of 0.56 μg/mL, and the ability to differentiate CD8a T cells from other components of the PBMC sample. In contrast, polymer <b>2f-CD8a</b> gave no Hg signal suggesting loss of Hg from the polymer during Ab conjugation or PBMC staining steps. Polymer <b>2d</b> represents an important step forward in the development of Hg mass tags for MC.