Universität Marburg, Fachbereich Chemie, Physikalische Chemie, Hans-Meerwein-Strasse, D-35032 Marburg, Germany.
Phys Chem Chem Phys. 2010 May 7;12(17):4345-50. doi: 10.1039/b923756a. Epub 2010 Feb 23.
Ultrathin carbon nanomembranes (CNM) have been tested as supports for both cryogenic high-resolution transmission electron microscopy (cryo-EM) as well as atomic force microscopy (AFM) of biological specimens. Purple membrane (PM) from Halobacterium salinarum, a 2-D crystalline monolayer of bacteriorhodopsin (BR) and lipids, was used for this study. Due to their low thickness of just 1.6 nm CNM add virtually no phase contrast to the transmission pattern. This is an important advantage over commonly used amorphous carbon support films which become instable below a thickness of approximately 20 nm. Moreover, the electrical conductivity of CNM can be tuned leading to conductive carbon nanomembranes (cCNM). cCNM support films were analyzed for the first time and were found to ideally meet all requirements of cryo-EM of insulating biological samples. A projection map of PM on cCNM at 4 A resolution has been calculated which proves that the structural integrity of biological samples is preserved up to the high-resolution range. CNM have also proven to be suitable supports for AFM analysis of biological samples. PM on CNM was imaged at molecular resolution and single molecule force spectra were recorded which show no differences compared to force spectra of PM obtained with other substrates. This is the first demonstration of a support film material which meets the requirements of both, cryo-EM and AFM, thus enabling comparative structural studies of biomolecular samples with unchanged sample-substrate interactions. Beyond high-resolution cryo-EM of biological samples, cCNM are attractive new substrates for other biophysical techniques which require conductive supports, i.e. scanning tunneling microscopy (STM) and electrostatic force microscopy (EFM).
超薄碳纳米膜（CNM）已被测试为支持低温高分辨率透射电子显微镜（cryo-EM）以及生物样本的原子力显微镜（AFM）。来自盐杆菌的紫色膜（PM），一种二维结晶的细菌视紫红质（BR）和脂质单层，被用于这项研究。由于其仅 1.6nm 的低厚度，CNM 几乎不会给透射模式增加任何相位对比。这是一个重要的优势，因为常用的无定形碳支撑膜在厚度约为 20nm 以下就变得不稳定。此外，CNM 的导电性可以被调整，从而产生导电碳纳米膜（cCNM）。cCNM 支撑膜首次被分析，被发现完全满足绝缘生物样本 cryo-EM 的所有要求。在 4A 分辨率下计算了 PM 在 cCNM 上的投影图，证明了生物样本的结构完整性一直保持在高分辨率范围内。CNM 也被证明是生物样本 AFM 分析的合适支撑。在 CNM 上对 PM 进行了分子分辨率成像，并记录了单分子力谱，与使用其他基底获得的 PM 的力谱相比没有差异。这是第一个满足 cryo-EM 和 AFM 两者要求的支撑膜材料的演示，从而能够对具有不变的样本-基底相互作用的生物分子样本进行比较结构研究。除了生物样本的高分辨率 cryo-EM 之外，cCNM 还是其他需要导电支撑的生物物理技术的有吸引力的新基底，例如扫描隧道显微镜（STM）和静电力显微镜（EFM）。
