Bio-AFM for combined light and atomic force imaging

 描述(由申请人提供)：我们建议获得一种原子力和光学显微镜的组合，这将对动态生物样本和软/湿材料研究(Bio-AFM)同样有用。The Bicope Resolve-Bio AFM(Bruker，Inc.)Head和AxioWatch Z1荧光显微镜(蔡司公司)我们要求的基座具有多功能的模块化设计，将原子力显微镜的亚分子分辨率成像和精确力测量与广泛的光学显微镜技术相结合，以同时进行相关的原子力显微镜和光学显微镜。这是一种高性能的Bio-AFM，由以下组件组成：1)Bio-AFM头，配备扩展z射程头、培养皿固定器和舞台加热器、二氧化碳控制、主动振动控制和声学封闭。2)安装AFM头的倒置荧光光学显微镜；提供自动化、衍射限制分辨率、广域荧光和亮场/相位成像能力，以及Piezo z驱动器、sCMOS机和分光镜，是利用AFM进行相关生物成像研究所需的。Bio-AFM及其所需配件将首先满足本提案中描述的NIH资助的研究项目的具体需求，然后满足更大的斯坦福大学和周围研究社区的预期需求。该仪器将实现创新的实验，允许在软材料、细胞和其他生物材料的表面进行高分辨率的力测量和测绘。这些力测量将与大分子、蛋白质和亚细胞结构相关联，因为细胞通过荧光、亮场和相衬光学成像来感知机械提示和环境变化并做出反应。这些成像实验的组合本来是不可能的，但将在多个学科和跨学科领域的重要健康相关研究课题上取得戏剧性和快速的进展，包括机械刺激感觉的分子和生物物理机制、再生医学研究的工程类器官培养、干细胞来源的心肌细胞收缩分析和家族性扩张型心肌病的建模、参与细胞分裂的蛋白质与微生物分裂中的力分布的关联以及T细胞信号通路的操纵以控制免疫介导性疾病。斯坦福大学的研究人员、学生和工作人员都很有资格--也渴望--进行这里描述的研究，使用Bio-AFM来推动这些和其他研究工作，为Bio-AFM开发和发布新的方法，并在共享成像设施内维护仪器，使更多的研究成为可能。

 DESCRIPTION (provided by applicant): We propose to acquire a combined atomic force and optical microscope that will be equally useful for dynamic biological sample and soft/wet material research (Bio-AFM). The BioScope Resolve-Bio AFM (Bruker, Inc.) head and AxioObserver Z1 epifluorescence microscope (Zeiss, Inc.) base we are requesting has a versatile, modular design that combines the sub-molecular resolution imaging and precision force measurements of AFM with a broad range of optical microscopy techniques for simultaneous, correlative AFM and optical microscopy. This is a high performance Bio-AFM consisting of the following components: 1) Bio-AFM head equipped with an extended z-range head, Petri dish holder and stage heater, CO2 control, active vibration control and acoustic enclosure. 2) Inverted epifluorescence optical microscope on which to mount the AFM head; provides automated, diffraction limited resolution, wide-field epifluorescence and brightfield /phase imaging capabilities with piezo z-drive, sCMOS camera, and splitting optics, needed for correlative biological imaging studies utilizing AFM. The Bio- AFM and its requested accessories will serve first the specific needs of the NIH funded research projects described in this proposal and then the anticipated needs of the greater Stanford University and surrounding research community. This instrument will enable innovative experiments that will allow high resolution force measurements and mapping over the surface of soft materials, cells and other biological material. These force measurements will be correlated with macromolecules, proteins and subcellular structures as cells sense and respond to mechanical cues and environmental changes via epifluorescence, brightfield and phase contrast optical imaging. These otherwise not possible combinations of imaging experiments will result in dramatic and rapid progress in important health-related research topics in a variety of disciplines and interdisciplinary areas including the molecular and biophysical mechanisms underlying sensation of mechanical stimuli, engineering organoid cultures for regenerative medicine research, analysis of stem cell derived cardiomyocyte contractility and modeling familial dilated cardiomyopathy, correlating proteins involved in cell division with force distribution in dividing microbes and the manipulatio of T cell signaling pathways to control immunologically-mediated diseases. Stanford researchers, students, and staff are well-qualified - and eager - to carry out the research described here, to use the Bio-AFM to further these and other research efforts, to develop and publish novel methodologies for Bio-AFM, and to maintain the instrument within a shared imaging facility that will enable additional research.