Copper-catalyzed click chemistry, an azide-alkyne cycloaddition reaction, is often used to label biomolecules on cell surfaces to study their functions. But it is not used in the cell cytoplasm because copper can be toxic to cells. A process that uses bulky ligands to stabilize and detoxify copper, enabling the reaction to be used inside living bacteria without hurting them, has now been developed by Jing Zhao of Nanjing University, in China; Peng Wu of Albert Einstein College of Medicine of Yeshiva University, in New York City; Peng R. Chen of Peking University, in China; and coworkers (Nat. Commun. 2014, DOI: 10.1038/ncomms5981). The researchers used tris(triazolylmethyl)amine-based ligands to corral copper as it helps fluorescently mark proteins with pH-dependent conformations in the cell cytoplasm and periplasm. This approach enabled them to determine pH gradients across the Escherichia coli cytoplasmic membrane. They also measured the E. coli transmembrane potential and determined the force required to move protons across the bacterial inner membrane under normal and acid-stress conditions.