Integrated approach to cell growth and recovery in silk fibroin scaffolds via a spin-down system

Silk fibroin scaffolds are a versatile platform for biomedical applications due to their biocompatibility and tunable properties. Successful clinical translation requires standardized production and characterization methods to ensure high reproducibility in cell seeding, growth profiling and recovery for downstream analysis. The intrinsic autofluorescence of silk and the limited diffusion of reagents through its porous structure present significant challenges for conventional assays, such as cell viability tests, DNA quantification, and optical imaging-based approaches. These assays are a requirement for validation procedures and characterization. In this study, we introduce a standardized protocol for efficiently assessing cell seeding and growth behavior. By analyzing the physicochemical properties of the silk sponge, we determined the optimal volumes required for silk swelling and cell seeding. Additionally, we developed a spin-down system that enables the application of endpoint assays while ensuring gentle cell recovery. We established and experimentally validated the relationship between silk sponge volume and the growth limitations of embedded cells. Overall, this study underscores the importance of a standardized procedure for efficient cell seeding and recovery, ultimately facilitating clinical translation.