Ling Zihan, Xing Yunhui, Jacome Emily Reinoso, Fok Shierly W, Ren Xi
Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, USA.
Bio Protoc. 2021 Feb 20;11(4):e3922. doi: 10.21769/BioProtoc.3922.
Decellularized extracellular matrix (ECM) biomaterials derived from native tissues and organs are widely used for tissue engineering and wound repair. To boost their regenerative potential, ECM biomaterials can be functionalized via the immobilization of bioactive molecules. To enable ECM functionalization in a chemoselective manner, we have recently reported an effective approach for labeling native organ ECM with the click chemistry-reactive azide ligand via physiologic post-translational glycosylation. Here, using the rat lung as a model, we provide a detailed protocol for and metabolic azide labeling of the native organ ECM using N-Azidoacetylgalactosamine-tetraacylated (AcGalNAz), together with procedures for decellularization and labeling characterization. Our approach enables specific and robust ECM labeling within three days or within one day during organ culture. The resulting ECM labeling remains stable following decellularization. With our approach, ECM biomaterials can be functionalized with desired alkyne-modified biomolecules, such as growth factors and glycosaminoglycans, for tissue engineering and regenerative applications.
源自天然组织和器官的去细胞细胞外基质（ECM）生物材料被广泛用于组织工程和伤口修复。为了提高其再生潜力，ECM生物材料可通过固定生物活性分子进行功能化。为了以化学选择性方式实现ECM功能化，我们最近报道了一种通过生理翻译后糖基化用点击化学反应性叠氮化物配体标记天然器官ECM的有效方法。在此，以大鼠肺为模型，我们提供了使用N-叠氮乙酰半乳糖胺-四酰化（AcGalNAz）对天然器官ECM进行代谢叠氮标记的详细方案，以及去细胞和标记表征的程序。我们的方法能够在三天内或在器官培养的一天内实现特异性和稳健的ECM标记。去细胞后，所得的ECM标记保持稳定。通过我们的方法，ECM生物材料可用所需的炔烃修饰生物分子（如生长因子和糖胺聚糖）进行功能化，用于组织工程和再生应用。
