Greve Frauke, Frerker Susanne, Bittermann Anne Greet, Burkhardt Claus, Hierlemann Andreas, Hall Heike
Physics Electronics Laboratory, Department of Physics, ETH Zurich, Switzerland.
Biomaterials. 2007 Dec;28(35):5246-58. doi: 10.1016/j.biomaterials.2007.08.010. Epub 2007 Sep 10.
Electrophysiological activities of neuronal networks can be recorded on microelectrode arrays (MEAs). This technique requires tight coupling between MEA-surfaces and cells. Therefore, this study investigated the interface between DRG neurons and MEA-surface materials after adsorption of neurite promoting proteins: laminin-111, fibronectin, L1Ig6 and poly-l-lysine. Moreover, substrate-induced effects on neuronal networks with time were analyzed. The thickness of adsorbed protein layers was found between approximately 1 nm for poly-l-lysine and approximately 80 nm for laminin-111 on platinum, gold and silicon nitride. The neuron-to-substrate interface was characterized by Scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and SEM after in situ focused-ion-beam milling demonstrating that the ventral cell membrane adhered inhomogeneously to laminin-111 or L1Ig6 surfaces. Tight areas of 20-30 nm and distant areas <1 microm alternated and even tightest areas did not correlate with the physical thickness of the protein layers. This study illustrates the difficulties to predict cell-to-material interfaces that contribute substantially to the success of in vitro or in vivo systems. Moreover, focused ion beam (FIB)/SEM is explored as a new technique to analyze such interfaces.
神经元网络的电生理活动可以在微电极阵列（MEA）上进行记录。该技术要求MEA表面与细胞紧密耦合。因此，本研究调查了在吸附神经突促进蛋白（层粘连蛋白-111、纤连蛋白、L1Ig6和聚-L-赖氨酸）后背根神经节（DRG）神经元与MEA表面材料之间的界面。此外，还分析了底物随时间对神经元网络的影响。在铂、金和氮化硅上，聚-L-赖氨酸吸附蛋白层的厚度约为1nm，层粘连蛋白-111吸附蛋白层的厚度约为80nm。通过扫描电子显微镜（SEM）和透射电子显微镜（TEM）对神经元与底物的界面进行了表征，原位聚焦离子束铣削后的SEM表明腹侧细胞膜与层粘连蛋白-111或L1Ig6表面的粘附不均匀。20-30nm的紧密区域和小于1微米的间隔区域交替出现，即使是最紧密的区域也与蛋白层的物理厚度无关。本研究说明了预测细胞与材料界面的困难，而这种界面对于体外或体内系统的成功至关重要。此外，聚焦离子束（FIB）/SEM被探索为一种分析此类界面的新技术。
