## Materials and Methods
### Characterization and Selection of the Prototype MCs  
#### 2.3.1 Selection of a Well-Performing Biodegradable MC Prototype: Cell Attachment Studies  
MCs were tested for their ability to allow the attachment of ASC52 telo cells under xeno- and serum-free conditions. To have a \"gold standard\" and positive control to compare our MCs' performance, we choose ProNectin-F MCs (Pall SoloHill, Port Washington, NY,  
Figure 1: Scanning electron microscopy (SEM) microphotographs to show the morphology of the _BR44_ microcarrier (MC). SEM examination of this carrier showed a good uniformity of shapes and sizes. _BR44_ is highly porous. Fold magnification: (**A**): 100\(\times\), scale bar 1 mm (**B,C**): 250\(\times\), scale bar 300 \(\upmu\)m (**D**): 500\(\times\), scale bar 200 \(\upmu\)m.  
USA). This commercial MC performed very well with our defined and other cell culture media [37]. The test used for this screening was as follows: (A) Preparation of the cells: The cells were detached from the cell culture vessel with TrypLE Select (after 2 min, incubation at 37 \({}^{\circ}\)C), washed with PBS supplemented with 1% human albumin, counted with Trypan Blue (Life Technologies, Thermo Fisher Scientific, Waltham, MA, USA) and resuspended in a culture medium at a concentration of 70,000 cells in 250 \(\mu\)L; (B) Preparation of the positive control MC: ProNectin-F MCs (PNF) were incubated in cell culture medium for one hour at 37 \({}^{\circ}\)C under agitation; C) Preparation of the test MCs: The different biodegradable MC prototypes manufactured by Sferalp were washed in PBS and then incubated in _UrSuppe_ for 60 min at 37 \({}^{\circ}\)C under agitation. Immediately before the tests, the MCs were resuspended in 250 \(\mu\)L of fresh _UrSuppe_ medium, resulting in a theoretical grow surface of 2.5 cm\({}^{2}\) for the ProNectin-F MC and 1 cm\({}^{2}\) for the _BR44_ prototypes. All of the screening tests were performed on low cell attachment plates (Ultra-Low 24 well Multiplate, Corning Inc., Corning, NY, USA). MC suspensions of 250 \(\mu\)L were transferred to each well, and the plate was gently mixed to distribute the MCs. Then, 250 \(\mu\)L of the ASC52 telo cell suspension was added. After gently mixing, the plate was placed in an incubator (37 \({}^{\circ}\)C, 5% CO\({}_{2}\)), and the cells were cultured for seven days. Every two to three days, 50% of the medium was removed and replaced with fresh, preheated _UrSuppe_ medium. After overnight incubation, it was usually possible to identify the low performing MCs since very few or no cells were attached to the carriers. The non-adherent cells tended to form multiple cellular aggregates (data not shown). Evaluation of the degree of colonization of the tested MCs was performed using phase-contrast microscopy, fluorescent microscopy, and SEM (see below). Of all the combinations of organic and biological polymers, shapes, porosities, and other tested parameters, the best combination that resulted in a carrier promoting cell attachment and proliferation comparable with a commercial product were _BR44_. Thus, this MC prototype was used for all project-specific experiments