5.8. Safranin O Staining

At the end of chondroinduction, the cell-seeded hDCB blocks were fixed in 4% PFA for 20 min at room temperature and washed several times in PBS. The formation of acidic proteoglycans by hMSCs on hDCB blocks was analyzed through staining with 0.1% Safranin O (Sigma-Aldrich) for 30 min. After 30 min of reaction, isopropanol was used to wash out the absorbed Safranin O for 20 min. The absorbance was detected at 540 nm with a Tecan Sunrise spectrophotometer according to a previous study [51]. Bone blocks without cells were used as background controls.