Trache Andreea, Meininger Gerald A
Texas A&M University System Health Science Center, Department of Medical Physiology, Cardiovascular Research Institute, College of Medicine, College Station, Texas 77843-1114, USA.
J Biomed Opt. 2005 Nov-Dec;10(6):064023. doi: 10.1117/1.2146963.
A novel hybrid imaging system is constructed integrating atomic force microscopy (AFM) with a combination of optical imaging techniques that offer high spatial resolution. The main application of this instrument (the NanoFluor microscope) is the study of mechanotransduction with an emphasis on extracellular matrix-integrin-cytoskeletal interactions and their role in the cellular responses to changes in external chemical and mechanical factors. The AFM allows the quantitative assessment of cytoskeletal changes, binding probability, adhesion forces, and micromechanical properties of the cells, while the optical imaging applications allow thin sectioning of the cell body at the coverslip-cell interface, permitting the study of focal adhesions using total internal reflection fluorescence (TIRF) and internal reflection microscopy (IRM). Combined AFM-optical imaging experiments show that mechanical stimulation at the apical surface of cells induces a force-generating cytoskeletal response, resulting in focal contact reorganization on the basal surface that can be monitored in real time. The NanoFluor system is also equipped with a novel mechanically aligned dual camera acquisition system for synthesized Forster resonance energy transfer (FRET). The integrated NanoFluor microscope system is described, including its characteristics, applications, and limitations.
构建了一种新型的混合成像系统，该系统将原子力显微镜（AFM）与提供高空间分辨率的光学成像技术相结合。该仪器（纳米荧光显微镜）的主要应用是研究机械转导，重点是细胞外基质-整合素-细胞骨架相互作用及其在细胞对外部化学和机械因素变化的反应中的作用。原子力显微镜可以对细胞骨架变化、结合概率、粘附力和细胞的微机械性能进行定量评估，而光学成像应用则可以在盖玻片-细胞界面处对细胞体进行薄切片，从而使用全内反射荧光（TIRF）和内反射显微镜（IRM）研究粘着斑。原子力显微镜与光学成像相结合的实验表明，细胞顶端表面的机械刺激会引发产生力的细胞骨架反应，导致基底表面的粘着斑重组，这一过程可以实时监测。纳米荧光系统还配备了一种用于合成荧光共振能量转移（FRET）的新型机械对准双相机采集系统。本文描述了集成的纳米荧光显微镜系统，包括其特点、应用和局限性。
