2.5. Live&Dead assay

Cell viability was detected by fluorescence live&dead assay at day 0 (right after bead preparation), 7, 14, and 21. Live cells were stained with calcein AM solution (cat.no. C1359; Sigma-Aldrich), while dead cells with cell membrane-impermeable Ethidium homodimer I solution (cat.no. E1903; Sigma-Aldrich). Cells were incubated for 30 min at RT and then washed in 1X PBS (Corning). Images were acquired at 4× magnification by a fluorescence microscope (Eclipse Ti Nikon Corporation, Tokyo, Japan). Images were acquired using the same settings (e.g., light intensity, exposure time, and gains). Signal intensity quantification was performed using ImageJ software (rel.1.52p National Institutes of Health, Bethesda, MD, USA), as previously described [32].