This study presents the 3D structure of vimentin intermediate filaments (VIFs) using cryo-focused ion-beam milling, cryo-electron microscopy, and tomography. VIFs are assembled into a modular, intertwined, and flexible helical structure composed of 40 α-helices in cross-section, organized into five protofibrils. The intrinsically disordered head domains form a fiber in the lumen of VIFs, while the intrinsically disordered tails form lateral connections between the protofibrils. The findings demonstrate how low-complexity protein domains can complement well-folded domains to construct a biopolymer with remarkable mechanical strength and stretchability. VIFs are expressed in mesenchymal cells and form dynamic, hyperelastic filament networks. They can be stretched up to 3.5 times their original length and are involved in various cytoskeletal processes, including cell shape maintenance, focal adhesion development, lamellipodia protrusion, stress fiber assembly, and signal transduction. VIFs are markers and regulators of the epithelial-mesenchymal transition and are involved in cancer progression and other pathophysiological conditions. At the molecular level, vimentin monomers form in-register, parallel dimers that are ~48 nm in length. These dimers contain a central coiled-coil rod domain composed of consecutive α-helical segments, connected by flexible linkers. The rod domain is flanked by intrinsically disordered head and tail domains. The early steps in VIF assembly involve the formation of elongated tetramers consisting of two antiparallel, staggered dimers, which are ~62 nm in length. The vimentin tetramer is the basic building block for subsequent filament assembly. The study used cryo-FIB milling and cryo-ET to analyze VIFs in situ. Subtomogram averaging and tomographic data revealed that VIFs are built from five protofibrils. Cryo-EM confirmed the helical symmetry of VIFs, and 3D structures of VIFs from human vimentin and tail-less vimentin were obtained. An atomic model of VIFs was constructed, showing a complex helical scaffold of intertwined vimentin tetramers that form protofibrils, surrounded by a central fiber composed of low-complexity head domains. The study also identified the position of the tail domains in the cryo-EM density map. The tail domains, despite their low sequence complexity, can condense into a stable structure in VIFs and establish connections between the protofibrils. The atomic model of VIFs revealed that the 2A–2B dimers shape the outer surface of VIFs, while the 1A–1B dimers coat the inner surface. The head domains form a fiber in the lumen ofThis study presents the 3D structure of vimentin intermediate filaments (VIFs) using cryo-focused ion-beam milling, cryo-electron microscopy, and tomography. VIFs are assembled into a modular, intertwined, and flexible helical structure composed of 40 α-helices in cross-section, organized into five protofibrils. The intrinsically disordered head domains form a fiber in the lumen of VIFs, while the intrinsically disordered tails form lateral connections between the protofibrils. The findings demonstrate how low-complexity protein domains can complement well-folded domains to construct a biopolymer with remarkable mechanical strength and stretchability. VIFs are expressed in mesenchymal cells and form dynamic, hyperelastic filament networks. They can be stretched up to 3.5 times their original length and are involved in various cytoskeletal processes, including cell shape maintenance, focal adhesion development, lamellipodia protrusion, stress fiber assembly, and signal transduction. VIFs are markers and regulators of the epithelial-mesenchymal transition and are involved in cancer progression and other pathophysiological conditions. At the molecular level, vimentin monomers form in-register, parallel dimers that are ~48 nm in length. These dimers contain a central coiled-coil rod domain composed of consecutive α-helical segments, connected by flexible linkers. The rod domain is flanked by intrinsically disordered head and tail domains. The early steps in VIF assembly involve the formation of elongated tetramers consisting of two antiparallel, staggered dimers, which are ~62 nm in length. The vimentin tetramer is the basic building block for subsequent filament assembly. The study used cryo-FIB milling and cryo-ET to analyze VIFs in situ. Subtomogram averaging and tomographic data revealed that VIFs are built from five protofibrils. Cryo-EM confirmed the helical symmetry of VIFs, and 3D structures of VIFs from human vimentin and tail-less vimentin were obtained. An atomic model of VIFs was constructed, showing a complex helical scaffold of intertwined vimentin tetramers that form protofibrils, surrounded by a central fiber composed of low-complexity head domains. The study also identified the position of the tail domains in the cryo-EM density map. The tail domains, despite their low sequence complexity, can condense into a stable structure in VIFs and establish connections between the protofibrils. The atomic model of VIFs revealed that the 2A–2B dimers shape the outer surface of VIFs, while the 1A–1B dimers coat the inner surface. The head domains form a fiber in the lumen of