The eukaryotic genome folds in 3D in a hierarchy of structures, including nucleosomes, chromatin fibers, loops, chromosomal domains (also called TADs), compartments and chromosome territories that are highly organized in order to allow for stable memory as well as for regulatory plasticity, depending on intrinsic and environmental cues. Our lab has provided evidence suggesting that the formation of TADs and chromatin loops can assist gene regulation, both in Drosophila and in mouse cells. Furthermore, cellular stress, such as replicative or oncogene-induced senescence, can induce a massive nuclear reorganization that can affect gene expression. However, the physical nature of compartments, TADs and loops remain elusive and single-cell studies are critically required to understand it. We characterized chromatin folding in single cells using super-resolution microscopy, revealing structural features inaccessible to cell-population analysis. TADs range from condensed and globular objects to stretched conformations. The physical insulation associated with their borders is variable between individual cells, yet chromatin intermingling is enriched within TADs compared to adjacent TADs in a large majority of cells. The spatial segregation of TADs is further exacerbated during cell differentiation. Favored interactions within TADs are regulated by cohesin and CTCF through distinct mechanisms. Furthermore, super-resolution imaging revealed that TADs are subdivided into discrete nanodomains. Altogether, these results provide a physical basis for the folding of individual chromosomes at the nanoscale. Our progress in these fields will be discussed.