The response of microorganisms to metal contamination of soils varies significantly from one investigation to another. One explanation is that metals are heterogeneously distributed at spatial scales relevant to microbes and that microoorganisms are able to avoid zones of intense contamination. This article aims to assess the microscale distribution of Cu in a vineyard soil. The spatial distribution of Cu was measured at two resolutions (0.3 mm and 20 μm) in thin sections of the surface 4 cm of undisturbed soil by electron microprobe and synchrotron X-ray microfluorescence spectroscopy. Bulk physicochemical analyses of Cu, pH, organic matter, texture, and mineralogy were performed. The results indicate that the Cu distribution is strongly heterogeneous at both scales of observation. Entire regions of the thin sections are virtually devoid of Cu, whereas highly localized “hotspots” have Cu signal intensities thousands of times higher than background. The distribution of Rb, or Al and Si, indicators of clay minerals, or Fe (iron (hydr)oxides), show that Cu is not preferentially associated with these mineral phases. Instead, Cu hotspots are associated with particulate organic matter. These observations suggest modification of current sampling protocols, and design of ecotoxicological experiments involving microorganisms, for contaminated soils.