Three dimensional (3D) bio-printing or direct writing technique has become a popular tool in tissue engineering applications that uses a computer-controlled process to deposit bio-ink for reproducing 3D tissue. Among multiple bio-printing modal, extrusion-based printing is capable of depositing diverse range of hydrogel materials and their compositions as bio-ink. Both acellular bio-ink and cell-laden bio-ink can be extruded by controlling the writing parameters to achieve high (>80%) cell survivability and density along with spatial precision and accuracy in 3D space. To increase cell viability and improve mechanical properties, nano-materials are often added in the bio-ink. However, the interplay between 3D bio-printing process parameters, solid fiber content and deposited fiber orientation has not been investigated yet. A novel cellulose based nano-fiber filled bio-ink (i.e. TEMPO nano fibrillated cellulose fiber) is developed and used in this research. The distribution of fiber is explored with respect to the 3D bio-printing process parameters such as nozzle diameter, applied pressure, fiber content and, alginate content. We found, fiber alignments has a very strong correlation with the deposition direction and about 70% fiber falls within 20 degree of the deposition direction.

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