2.2 Evaluation of decellularization
2.2.1 Histological and immunofluorescent evaluation
For confirmation of decellularization, DP samples were fixed in 4% paraformaldehyde for 24 h and embedded in paraffin. After dehydration and paraffin embedding, the DP samples were cut into 5-μm-thick sections for histological examination using hematoxylin and eosin (H&E) staining. The stained sections were visualized and photographed on an optical microscope (Leica, DM3000, Germany). Residual nuclei within the DP were observed by DAPI staining viewed on an Olympus BX53 microscope (Olympus, Japan).
2.2.2 Quantification of DNA, glycosaminoglycan (GAG) and collagen content
The DNA content in DP samples was determined using a Genomic DNA Extraction Kit (Takara, China) following the manufacturer’s instructions. The GAG content in DP samples was evaluated using a DMMB Colorimetry Kit (GenMed, USA). A hydroxyproline assay kit (Abcam, USA) was utilized to quantify the collagen content of DP samples according to the manufacturer’s instructions.