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International Journal of Bioprinting Automated bioink mixer improves bioprinting

Figure 4. Hydrogel mixing at different conditions using the automated device. Hydrogel mixing was preliminarily explored for different alginate
concentrations (A), mixing speeds (B), mixture volume (C), and volume ratios (methylene blue (MB) solution volume : alginate hydrogel volume) (D).
Alginate concentration of 6% was tested (B, C), while various concentrations were used (D; Table S1 in Supplementary File). The mixing effect was assessed
at regular intervals of exchanges during the process. The larger numbers indicate the better blending effect, and the black thick lines mark the critical point
where the hydrogel reached a homogeneous state under the given conditions.

The volumes of the different components are also likely the system before mixing. Both of them can facilitate the
to make a difference for the mixing behavior. Therefore, mixing effect.
the mixing process was further examined using different
total volumes ranging from 1 mL to 3 mL at a speed of 3.3. Impact of mixing conditions on cell distribution
10 mm/s. As illustrated in Figure 4C, the number of and viability
exchanges required to achieve homogeneity increased Cell distribution and viability are greatly influenced by the
mixing process, in addition to the bioprinting conditions.
with the volume of the mixed components due to their To investigate how the mixing process impacts bioink
increased longitudinal dimensions. homogeneity and cell status, a systematic exploration
Additionally, experiments were conducted with was carried out with different mixing parameters. The
different volume ratios (methylene blue volume to alginate two fundamental parameters for mixing bioink using a
hydrogel volume) with a final alginate concentration of syringe coupler are the mixing speed and the number
3% at 10 mm/s. As shown in Figure 4D, ratios of 1:3 and of exchanges. As described for the manual mixing
1:1 required the fewest number of exchanges to obtain experiments outlined above, a hydrogel consisting of
homogeneous mixtures. For the case of 1:3 ratio, a lower 6% alginate and a suspension of HEK293-GFP cells
initial alginate concentration of 4% was required to obtain were used.
a final value of 3%. This is lower than other ratios, with Initially, the components were mixed for 100
initial concentrations ranging from 4.5% to 12%, and exchanges at different speeds ranging from 5 mm/s to
therefore, the initial viscosity of the hydrogel component 50 mm/s. As can be seen in the fluorescence microscopy
was the lowest. When methylene blue solution and images shown in Figure 5A, no substantial differences
hydrogel were mixed in equal volumes, both components of cell distribution in samples from different positions
had the maximum probability of overall distribution in were observed after 100 exchanges for any of the tested

Volume 10 Issue 2 (2024) 387 doi: 10.36922/ijb.1974