MultimodalProfiling of Iron Heterogeneity at theNanoscale

Life is inherently heterogeneous, and visualizing this heterogeneity in the spatial distribution of biometals offers valuable insights into various biological processes. While biometal mapping provides superior spatial resolution compared to other bioanalytical techniques, it alone cannot fully explain the functional roles of biometals in health and disease. In this study, we introduced a novel method using specially designed sample grids to facilitate beam-, X-ray-, and ion-beam-based imaging of biometals on the same tissue section. This innovative approach aligns and integrates nanoscale-resolved iron profiles across spatial, chemical, and isotopic dimensions. By combining these analyses, we achieved unprecedented spatial resolution and detail, revealing the complex regulatory framework of iron homeostasis in liver tissues following iron overload. These findings demonstrate that enhancing both the information content and spatial resolution of biometal analysis can overcome current limitations, providing new insights into the molecular mechanisms underlying biometal functions.