Liu Yuanfang, Wang Shaopeng
1024S Innovation Way, Stillwater, OK 74074, USA.
Colloids Surf B Biointerfaces. 2007 Jul 1;58(1):8-13. doi: 10.1016/j.colsurfb.2006.08.014. Epub 2006 Aug 22.
The measurement of local oxygen level in 3D cell culture is desired but remains as a challenge problem. We developed a 3D cell scaffold with luminescence-based oxygen sensing capability that opens the possibility of 3D mapping of oxygen level during cell growth. Hydrogel inverted opal scaffold was prepared by photo-polymerization of poly(2-hydroxyethyl methacrylate (pHEMA) and poly(methacryloyloxy)ethyl-trimethylammonium chloride (pMEATAC) monomer using close-packed bead assembly as template. Tris(4,7-diphenyl-1,10-phenanthroline)ruthenium chloride (Ru(dpp)(3)), was coated on the pHEMA-pMEATAC 3D scaffolds by layer-by-layer (LBL) assembly. pHEMA-pMEATAC copolymer was coated on top of the Ru(dpp)(3) layer as a protection layer. The fluorescence emission of Ru(dpp)(3) can be dynamically quenched by oxygen. By measuring the emission intensity of the scaffold, the local oxygen level can be monitored. The hydrogel scaffolds are transparent, and thus 3D fluorescence intensity can be mapped by confocal microscopy. Human bone marrow stromal cells HS-5 were successfully cultured on the oxygen sensing scaffold, and the observed Ru(dpp)(3) emission intensity from the scaffold was stronger in cell rich area, which indicates a lower oxygen level due to the consumption of the cells.
在三维细胞培养中测量局部氧水平是人们所期望的，但仍然是一个具有挑战性的问题。我们开发了一种具有基于发光的氧传感能力的三维细胞支架，这为细胞生长过程中氧水平的三维映射提供了可能性。通过使用密排珠组装作为模板，对聚（甲基丙烯酸2-羟乙酯）（pHEMA）和聚（甲基丙烯酰氧基）乙基三甲基氯化铵（pMEATAC）单体进行光聚合制备水凝胶反蛋白石支架。通过逐层（LBL）组装将三（4,7-二苯基-1,10-菲咯啉）氯化钌（Ru(dpp)(3)）涂覆在pHEMA-pMEATAC三维支架上。将pHEMA-pMEATAC共聚物作为保护层涂覆在Ru(dpp)(3)层的顶部。Ru(dpp)(3)的荧光发射可以被氧动态猝灭。通过测量支架的发射强度，可以监测局部氧水平。水凝胶支架是透明的，因此可以通过共聚焦显微镜对三维荧光强度进行映射。人骨髓基质细胞HS-5成功地在氧传感支架上培养，并且在富含细胞的区域观察到来自支架的Ru(dpp)(3)发射强度更强，这表明由于细胞消耗导致氧水平较低。
