Real-Time Intracellular Mg2+ Signaling and Wave Propagation by Subdiffraction-Limit Super-Resolution Microscopy

Abstract

Magnesium ion (Mg2+) is an important intracellular component that plays a fundamental role in many cellular processes. The super-localization of intracellular Mg2+ signaling and wave propagation in a single cell with subdiffraction-limit image resolution is demonstrated for the first time by direct stochastic optical reconstruction microscopy (dSTORM). Mag-fluo-4-AM, an intracellular Mg2+ fluorescent indicator dye, is used for the fluorescence imaging of intracellular Mg2+ in human embryonic kidney 293 (HEK-293) cells as a model. Mag-fluo-4-AM is photoswitched between its fluorescent and dark states and the resulting super-resolution cell image reveals uniform and specific labeling of Mg2+ with thousands of fluorophores per cell and localization precision of 21 ± 1 nm. Furthermore, the real-time dynamics of Mg2+ wave propagation are studied by dSTORM. The sparks and waves show random temporal propagation patterns in nonhomogeneous substructures with 0.2 s time intervals. Thus, dSTORM allows characterization of intracellular processes at sub-nanometre spatial resolution with high temporal resolution.