An Improved Atomic Force Microscope for Biomedical Applications: Deep AFM for lar

描述（由申请人提供）：需要一种新工具来探索有助于组织机械性能的纳米级机制。这些纳米机制对于理解骨脆性、椎间盘退变、骨关节炎和其他疾病具有关键重要性。原子力显微镜（AFM）具有成像和操纵纳米结构的能力，已成为这一领域的有力工具。但它在研究组织样品时有一个主要的局限性：大多数组织样品太粗糙，无法用AFM杠杆成像。在这里，我们提出了一种新的深度AFM探针，使一个垂直的方法深入到组织样本的地形超越杠杆。拟议研究的目标是建立新一代原子力显微镜的第一个原型，深度AFM，这将增加AFM的成像范围至少一个数量级，然后使用这些深度AFM来探索组织结构和纳米力学，目的是以足够的细节水平了解组织变性中涉及的分子和纳米级过程，以告知新疗法由于许多潜在的应用，拟议工作的整体影响和相关性是广泛的，但是，由于我们目前在骨诊断方面的进展和工作，我们将主要关注骨的纳米级断裂力学。我们提出了三个相关的目标，包括这类新的AFM的开发和表征，以及它们在临床相关骨组织样本中的应用。具体目标1是开发用于超大规模扫描和纳米力学的Deep AFM I。它将使浸没在缓冲液中的骨中的裂纹扩展成像以及空间分辨力谱、纳米操纵和压痕能够测量局部纳米力学性能。具体目标2是开发用于高分辨率、大规模扫描和纳米力学的Deep AFM II。Deep AFM II的更高分辨率将能够以与扫描电子显微镜相当的分辨率对骨折裂纹的纳米级起源进行成像，但无需将样品从缓冲液中取出。因此，将有可能对在裂纹生长过程的中间阶段发生的纳米级过程进行成像。它也将有可能进行空间分辨力谱，纳米操纵和压痕测量局部纳米机械性能。具体目标3是使用深度AFM来推进我们对骨折的纳米机制和降低骨折风险的方法的理解。有了Deep AFM，我们可以通过了解抵抗骨折的分子和纳米级机制，继续朝着临床降低骨折风险的长期目标迈进。 公共卫生相关性：骨脆性、椎间盘退变、癌症、动脉粥样硬化、骨关节炎和蛀牙都涉及纳米级组织机械特性的变化。拟议的深原子力显微镜，深原子力显微镜，旨在研究这些变化，目的是了解如何防止甚至逆转一般组织，特别是骨骼的不良变化。这项工作将与我们正在进行的与医生合作诊断患者骨组织变性引起的骨脆性产生协同作用。

DESCRIPTION (provided by applicant): A new tool is necessary to explore the nanoscale mechanisms that contribute to tissue mechanical properties. These nanoscale mechanisms are of key importance in understanding bone fragility, intervertebral disc degeneration, osteoarthritis, and other diseases. The Atomic Force Microscope, AFM, with its ability to both image and manipulate nanostructures, has been a powerful tool in this area. But it has a major limitation for studying tissue samples: most tissue samples are too rough to be imaged with AFM cantilevers. Here we propose to move beyond cantilevers to a novel Deep AFM probe that enables a vertical approach deep into the topography of tissue samples. The objectives of the proposed research are to build the first prototypes of a new generation of Atomic Force Microscopes, Deep AFMs, that will increase imaging range of AFMs by at least one order of magnitude, and then to use these Deep AFMs to explore tissue structures and nanomechanics with a goal of understanding the molecules and nanoscale processes involved in tissue degeneration at a sufficient level of detail to inform development of new therapies. The overall impact and relevance of the proposed work is broad due to the numerous potential applications, however, because of our current progress and work on bone diagnostics, we will focus primarily on the nanoscale fracture mechanics of bone. We propose three related aims that include both the development and characterization of this new class of AFMs as well as their application in clinically relevant bone tissue samples. Specific Aim 1 is to develop Deep AFM I for very large scale scanning and nanomechanics. It will enable imaging of crack propagation in bone submerged in buffer as well as spatially resolved force spectroscopy, nanomanipulation and indentation to measure local nanomechanical properties. Specific Aim 2 is to develop Deep AFM II for high resolution, large scale scanning and nanomechanics. The higher resolution of Deep AFM II will enable imaging the nanoscale origin of bone fracture cracks with resolution comparable to Scanning Electron Microscopy, but without ever removing the sample from buffer. Thus it will be possible to image nanoscale processes that occur at intermediate stages of the crack growth process. It will also be possible to perform spatially resolved force spectroscopy, nanomanipulation and indentation to measure local nanomechanical properties. Specific Aim 3 is to use Deep AFM to move forward in our understanding of the nanoscale mechanisms of bone fracture and ways to reduce bone fracture risk. With Deep AFM, we can continue to move toward a long term goal of clinically decreasing the component of bone fracture risk by understanding the molecules and nanoscale mechanisms that resist bone fracture. PUBLIC HEALTH RELEVANCE: Bone fragility, intervertebral disc degeneration, cancer, atherosclerosis, osteoarthritis, and tooth decay all involve changes in tissue mechanical properties at the nanoscale. The proposed Deep Atomic Force Microscope, Deep AFM, is designed to investigate these changes with the goal of understanding how to prevent and even reverse undesirable changes for tissues in general and bone in particular. This work will be synergistic with our ongoing collaboration with physicians on diagnosing bone fragility due to bone tissue degeneration in patients.