Printing Photocrosslinkable Bioinks

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Bioprinting processes often make use of natural materials such as collagen, alginate, gelatin, and cellulose. These materials usually are preferred over synthetic ones for bioink formulation since they have higher biocompatibility. However, incorporating natural polymers into bioinks is a recurrent challenge due to their lack of mechanical and structural stability, leading to poor printability and deficient shape fidelity of the final product [1,2].

A solution to this disadvantage is inducing gelation through photocrosslinking mechanisms, which rely on photoinitiators to promote reactions to light stimuli. In the photocrosslinking process, the most widespread technique to increase stiffness on bioinks is irradiation with ultraviolet (UV) light [1,2]. Figure 1 shows different ways to perform the bioink photocuring process when using extrusion-based bioprinters: before (pre-crosslink), after (post-crosslink), or during (in-situ-crosslink) extrusion [3].

Figure 1: Methods for photocrosslinking bioinks when performing extrusion-based bioprinting [3].

Choosing the right photoinitiator is a critical step in photocrosslinking-based bioprinting as the initiator’s absorption peak dictates the wavelength of light that must be applied to the bioink. Even considering UV light’s wide use, some studies have shown that this type of light fails to penetrate deep inside the scaffold, crosslinking only the printed construct’s surface layer [4,5]. However, blue light operates at a wavelength quite close to the UVA band, which seems not to affect cells as severely as UV light. Some researchers have been working on developing visible light-based photocrosslinking of bioinks in order to mitigate cytotoxic effects after exposure to light.

Lim et al. demonstrated this comparison in a study using the conventional UV method and a new visible light-initiating method, with the purpose to assess the differences between the final product when these different approaches are used [6].

This study described the development and characterization of an alternative bioink and photoinitiator method suitable for biofabrication featuring an improved printing window and a favorable crosslinking method. In comparison to the conventional UV system, an increase in cell cytocompatibility was observed when the new visible light system (Ruthenium/ Sodium persulfate, Ru/SPS) system was used. This improvement was evidenced by higher cell viability at high Ru/SPS photoinitiator concentrations or visible light irradiation intensities [6].

Through a different technique, Galarraga et al. [7] were able to crosslink a norbornene-modified hyaluronic acid hydrogel containing an UV-light based photoinitiator via in situ crosslinking. This technique permits the printing of non-viscous, photocrosslinkable bioinks via the direct-curing of the bioink with light through a photo permeable capillary before the deposition. The printed constructs turn out to be cytocompatible at the final printing process, with high cell viability and homogenous distribution of mesenchymal stromal cells (MSCs). The focus of this study was on the repair of focal defects in articular cartilage. However, it can be easily expanded to other cell types and tissue of interest, not only for clinical applications of tissue repair but also for in vitro models or drug screening [7].

Figure 2: Representative multi-layered constructs printed via in situ crosslinking. Left: Schematic of in situ crosslinking method and Right: CAD design and representative image of a printed construct for designs of (a) a model femoral condyle or (b) a disc [7].


1. Serna, J. A. et al. Formulation and Characterization of a SIS-Based Photocrosslinkable Bioink. Polymers 11, (2019).

2. Hong, B. M., Park, S. A. & Park, W. H. Effect of photoinitiator on chain degradation of hyaluronic acid. Biomater Res 23, 21 (2019).

3. Zheng, Z. et al. Visible Light-Induced 3D Bioprinting Technologies and Corresponding Bioink Materials for Tissue Engineering: A Review. Engineering (2020) doi:10.1016/j.eng.2020.05.021.

4. Wang, Z. et al. A simple and high-resolution stereolithography-based 3D bioprinting system using visible light crosslinkable bioinks. Biofabrication 7, 045009 (2015).

5. Knowlton, S., Yenilmez, B., Anand, S. & Tasoglu, S. Photocrosslinking-based bioprinting: Examining crosslinking schemes. Bioprinting 5, 10–18 (2017).

6. Lim, K. S. et al. New Visible-Light Photoinitiating System for Improved Print Fidelity in Gelatin-Based Bioinks. ACS Biomater Sci Eng 2, 1752–1762 (2016).

7. Galarraga, J. H., Kwon, M. Y. & Burdick, J. A. 3D bioprinting via an in situ crosslinking technique towards engineering cartilage tissue. Sci. Rep. 9, 19987 (2019).

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