Assessing metal ion transporting activity of ZIPs: Intracellular zinc and iron detection

Zrt/Irt-like proteins (ZIPs or SLC39A) are a large family of metal ion transporters mainly responsible for zinc uptake. Some ZIPs have been shown to specifically transport zinc, whereas others have broader substrate specificity in divalent metal ion trafficking, notably those of zinc and iron ions. Measuring intracellular zinc and iron levels helps assess their molecular and physiological activities. This chapter presents step-by-step methods for evaluating intracellular metal ion concentrations, including direct measurement using inductively coupled plasma-mass spectrometry (ICP-MS), chemical staining, fluorescent probes, and indirect reporter assays such as activity analysis of enzymes whose activities are dependent on metal ion availability.