CARBON NANOTUBES AS KNIVES FOR CUTTING VITREOUS ICE

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。埋藏在无定形冰中的冷冻水化生物样品的层析重建是研究原始状态下细胞结构的一种强有力的方法。然而，很少有生物样品沿着束轴小于0.5微米，薄到足以用300keV电子观察它们的整体。因此，重要的是要开发出冰冻细胞切片的方法。在良好的冷冻-超微切割机中的钻石刀可以切割所需的部分，而不会将它们加热到无定形冰将结晶的温度，但稳定的钻石刀很厚(刀刃上的角度为25-35度)，因此无定形冰通常在切割过程中被压缩和急剧弯曲。为了解决这些问题，我们正在研究使用碳纳米管(CNT)作为设备，这种设备可以用作紧线，从玻璃化的生物材料块上切下薄片。单壁碳纳米管的半径约为1 nm，小于锋利的钻石刀边缘的曲率。碳纳米管可以有几毫米长，而且众所周知是坚固的，但相对于无定形冰的强度还有待确定。我们在扫描电子显微镜中观察了商业上获得的和国产的碳纳米管，这些碳纳米管通过显微操作从相邻的碳纳米管中分离出来。我们将它们拉伸到一条细长的钨丝环上，并开发出一种在碳纳米管和钨之间形成牢固结合的“焊接”工艺。我们正在努力简化这一过程，以便它可以用于制造大量有用的基于碳纳米管的切割设备。我们还在试验可以测量玻璃体冰的硬度和粘度的设备，这样我们就可以了解我们计划进行的显微切割的物理要求。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Tomographic reconstruction of frozen-hydrated biological samples, embedded in amorphous ice, is a powerful method for the study of cell structure in its initially native condition. However, few biological samples are less than 0.5 micrometers along the beam axis, thin enough to be viewed in their entirety with 300 KeV electrons. It is therefore important to develop methods for slicing cells while they are frozen. A diamond knife in a good cryo-ultramicrotome can cut the desired sections without warming them to temperatures at which amorphous ice will crystallize, but stable diamond knives are thick (the angle included at the knife edge is 25-35 degrees), so the amorphous ice is usually compressed and sharply bent by the cutting process. To solve these problems we are investigating the use of carbon nanotubes (CNTs) as devices that could be used as tight wires to sever thin slices from a block of vitrified biological material. Single walled CNTs have a radius of about 1 nm, less then the curvature at the edge of a sharp diamond knife. CNTs can be millimeters long and are known to be strong, but how strong relative to amorphous ice remains to be determined. We have examined commercially obtained and homegrown CNTs in a scanning electron microscope, disentangled from their neighbors by micromanipulation. We stretch them over a thin loop of tungsten wire and have developed a "welding" process that forms a strong bond between the CNT and the tungsten. We are working to streamline this process so it can be used to make useful numbers of CNT-based cutting devices. We are also experimenting with devices that can measure the hardness and viscosity of vitreous ice, so we can understand the physical requirements for the microtomy we plan.