## 3 Results  
We extracted virtual models of human ossicles from a high-quality open-source database of animal ossicle chains (Figure 1A), which was imported in the bioprinter's BioCAD program. The printable files are shown here at single-layer level (Figure 1B), layer-filling trajectory (Figure 1C), and as the progression of several layers on top of each other (Figure 1D) of an incus print. For printing, we first used the plottable CPC of calcium phosphate/hydroxyapatite paste. Initially, a flat base was required to support this soft material, which was obtained by omitting the bottom layers of the constructs. In this way, we printed in triplicate the malleus and incus on glass slides, showing that the constructs retained the desired structure and external texture (Figure 2A). The models were cured by heating at 37 \({}^{\circ}\)C in 100% humidity for five days for hardening, as recommended by the CPC provider. Due to this treatment, the material became solid, with no discernible changes in structure or exterior texture of the printed ossicle models (Figure 2B). The stages was the most challenging to print, as it is the smallest of the three ossicles. For this reason, the stages model was also made flat on the bottom and printed directly on a glass plate (Figure 2C). Despite its small size, all four anatomical characteristics of the stages still appeared visible: the head, anterior crus, posterior crus, and footplate. Due to the small volume of material, the stages model only required three days of curing at 37 \({}^{\circ}\)C in 100% humidity. The model was then strong enough to be removed from the support without fracturing. Additionally, there were no discernable changes to the surface exterior after curing (Figure 2D). To test the reproducibility of the procedure with this material, we printed 10 incurs models and solidified them as before, then assessed their size both by visual inspection and by mass. We found the models to be reproducible, although minor differences in the  
Figure 1: **CAD of adult human ossicles and bioprinter toolpath for their generation**. (**A**) 3D model of the middle ear ossicles (malleus, blue; incus, green; and stapes, purple) [12]. (**B**) Single-layer toolpath created and extracted with the BioCAD program from an incus model. (**C**) Total filling trajectory of an incus print, as generated by the program. (**D**) Snapshots of layer progression for the same ossicle. Bar, 1 mm. Figure 2: **Ossicle models printed on a solid support. (A) Triplicate prints with CPC paste of the incus and malleus, left and right, respectively (imaged before curing). (B) Printed malleus and incus after curing for five days. (C) Two printed stapes models before curing. (D) Printed stapes model after curing for two days. (E) Ten incus prints after curing, showing replicability in size (see also Table S1) and texture, with only minor differences between the individual objects. Bars, 2 mm. **  
external texture could be occasionally observed, and with a relative standard variation of the weight within 6% (Table S1). To generate whole constructs by direct printing on a substrate, the CADs of incus and malleus were split in two, and the individual halves were printed facedown directly on a glass plate (Figure 3A,B). The prints were cured under the same conditions as previously described and assembled by adhesion with a superglue, re-forming the full ossicle models (Figure 3C,D). As a more straightforward way to generate whole ossicle models, we also printed the models by an 'embedding' method using Carbopol as the supporting hydrogel [17]. The shape of the printed objects was properly maintained within the Carbopol hydrogel bath, with the additional advantage of easy visual monitoring during and after printing due to its robustness and transparency. The objects were printed within the wells of a 12-well plate (Figure 4A) and also in a gel support directly placed on a glass slide for high-temperature curing (Figure 4B). This method allowed the easier extraction and solidification of the models as well (Figure 4C).