Bio-based resource utilization in different forms has been driven by societal, industrial and academic research interests towards the development of “green”, sustainable materials from renewable sources. Within this context, exploiting biomass from different industrial residues is further advantageous from an environmental and economic point of view, leading to minimization of residues by means of waste treatment and to the development of high-addedvalue- products. Breaking down the cell wall structure to its smallest structural components is one means of turning bio-based residues into high-value products, leaving us with nanofibers. The aim of this work has been to understand how these nanofibers can be liberated from various cellulosic sources using mechanical fibrillation and how they can be assembled into functional hydrogels. The production of bio-based nanofibers as a sustainable bio-based material is in the early stages of commercialization and considerable research has been devoted to explore different methods of reaching nanoscale. However, the extraction process by chemical and/or mechanical means is still associated with a relatively high energy demand and/or cost. These are key obstacles for use of the material in a wide range of applications. Another challenge is that methods to characterize nanofiber dimensions are still being developed, with few options available as online measurements for assessing the degree of fibrillation. Allowing for assessment during the fibrillation process would enable not only optimization towards a more energy efficient fibrillation, but also matching of the nanofiber quality to its intended function, since different applications will require widely different nanofiber qualities. Energy-efficient fibrillation and scalability from industrial residues were explored using upscalable ultrafine grinding processes. Nanofibers from various industrial bio-residues and wood were prepared and characterized, including the development of a method for evaluation of the fibrillation process online via viscosity measurements as an indication of the degree of fibrillation down to nanoscale. Furthermore, the correlation of viscosity to that of the strength of the nanopapers (dried fiber networks) was evaluated for the different raw materials. Switchable ionic liquids (SIL) were tested as a green pretreatment for delignification, without bleaching of wood prior to fibrillation, with the aim to preserve the low environmental impact that the raw material source offers. In order to employ the hydrophilic nature and strong network formation ability of the fibrillated nanofibers, they were utilized in the preparation of functional biomaterials in the form of hydrogels. Firstly, brewer’s spent grain nanofibers were used to promote and reinforce hydrogel formation of lignin-containing arabinoxylan, resulting in a hydrogel completely derived from barley residues. In addition, alginate-rich seaweed nanofibers from the stipe (stem-like part of the seaweed) were used directly after fibrillation as an ink and hydrogels were formed via 3D printing.