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Laser-based bioprinting allows clearer view of cell dynamics

23 Oct 2019

TU Wien two-photon polymerization platform embeds cells into bioinks.

Allowing cells and tissues to develop ex vivo on 3D polymer frameworks has become a vital part of studying biological processes.

Additive techniques based on 3D printing are a key aspect of both manufacturing the frameworks and successfully embedding the cells within them, but the materials involved need to be carefully selected and the printing operation must not damage the cells.

A project at Vienna's TU Wien has now developed a high-resolution bioprinting process that allows cells to be embedded within a 3D matrix with micrometer precision, and at a printing speed of one meter per second, said to be orders of magnitude faster than previously possible. The work was published in Advanced Healthcare Materials.

"The behavior of a cell depends crucially on the mechanical, chemical and geometric properties of its environment," said Aleksandr Ovsianikov, head of the 3D Printing and Biofabrication group at TU Wien. "The structures in which the cells are embedded must be permeable to nutrients so that the cells can survive and multiply. But it is also important whether the structures are stiff or flexible, whether they are stable or degrade over time."

The breakthrough involves the use of two-photon polymerization (2PP), whereby the gels used to build the 3D matrix solidify through the absorption of two photons of required wavelength. This requires a higher power laser than conventional polymerization, but in return is capable of spatial resolution in the sub-micrometer range.

Two-photon processes have been used in lithography processes for some time as a means to create small features or photoresists in a photosensitive material, and TU Wien has now taken the concept into the bioprinting arena.

Tailor-made tissues

"Due to the non-linear nature of 2PP process, it is possible to produce structures directly within the volume of the sample," noted the team in its paper. "This eliminates the need for a layer-by layer deposition, or the addition of absorbers in order to limit penetration depth, as in stereolithography and digital light processing."

These advances have already spawned a TU Wien spin-out, UpNano, formed in 2018 specifically to commercialize high-resolution 3D printing systems based on a two-photon polymerization operation through the company's NanoOne platform.

In the TU Wien bioink trials, direct embedding of mouse fibroblast cells into gelatin–norbornene bioinks was performed via 2PP, using a commercial femtosecond 720 nm laser source. The study is thought to be the first systematic study of embedding cells in hydrogel constructs by 2PP technology at relatively high throughput.

"The high reactivity of the thiolene system allows 2PP processing of cell-containing materials at remarkably high scanning speeds of 1000 millimeters per second, placing this technology in the domain of bioprinting," commented the team. The 3D-printed constructs supported cell adhesion and migration as desired, and the photosensitive bioink enabled high-definition bioprinting of well-defined constructs for long-term cell culture studies.

"Using these 3D scaffolds, it is possible to investigate the behavior of cells with previously unattainable accuracy. It is possible to study the spread of diseases," said Ovsianikov. "And if stem cells are used, it is even possible to produce tailor-made tissue in this way".

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