Spinning disk confocal laserscanning microscope
转盘共焦激光扫描显微镜
基本信息
- 批准号:417424858
- 负责人:
- 金额:--
- 依托单位:
- 依托单位国家:德国
- 项目类别:Major Research Instrumentation
- 财政年份:2019
- 资助国家:德国
- 起止时间:2018-12-31 至 无数据
- 项目状态:未结题
- 来源:
- 关键词:
项目摘要
The microscope to be acquired should enable the live cell imaging of single cells but also of cells within a three-dimensional tissue. It is intended to be used by a number of groups which work together in the newly founded Zentrum für Pharmaverfahrenstechnik (PVZ , Center of Pharmaceutical Engineering) of the TU Braunschweig. Thus, its measuring principle should allow for the flexible adaption to the diverse requirements of the various cellular models employed by these groups. In the Z-dimension the resolution should be sufficient to resolve multiple planes per cell, while in the X/Y-dimension a high resolution but not necessarily super-resolution is desirable. Overall, the characteristics should provide a balance between spatial resolution and temporal resolution. Frame rates of more than 10 Hz should be achievable (e.g. to document changes in the cytosolic calcium together with the mobility of mitochondria or secretory granules) without requiring a strength of fluorescence excitation which causes photobleaching or phototoxicity. The avoidance of toxicity is also relevant for long-term measurements of continuously perifused cells or tissues. These features should come along with a depth of penetration that is sufficient to handle 3D model organs (organoids). These requirements are met by the spinning disk confocal microscopy. In contrast to the "conventional" variant of the confocal laser scanning microscope which sequentially excites the fluorescence of one image point at a time, the spinning disk (Nipkow disk) variant excites a multitude of fluorescent image points at a time by virtue of a rotating disk containing a spiral pattern of pinholes. The image generation is two-dimensional from the beginning on, permitting the use of a camera chip as the detecting element. Originally, the spinning disk principle suffered from the low light transmission. Two developments have contributed to make the spinning disk an instrument for high sensitivity fluorescence detection. First, the excitation light is focused on the pinhole pattern by a corresponding pattern of microlenses in a parallel second disc. Second, the detection efficiency was greatly improved by the recent evolution of sCMOS cameras, which combine a very high sensitivity with a small pixel size. Since each point in the object plane is scanned multiple times during image generation (depending on the rotation velocity of the disk), the excitation energy per image point and time is much lower than in the conventional laser scanning microscopes. This enables long-term observations without phototoxicity and/or high rates of image acquisition. The latter feature can be used for fast z-stacking within a cell and thus enables the description of functional changes in 3D specimen without aliasing. Taken together, the spinning disk principle in its current state of development has resulted in a very versatile method to describe functional changes with high spatial and temporal resolution.
要获取的显微镜应该能够对单个细胞以及三维组织内的细胞进行活细胞成像。它旨在供在布伦瑞克工业大学新成立的 Zentrum für Pharmaverfahrenstechnik(PVZ,制药工程中心)中合作的多个小组使用。因此,其测量原理应该能够灵活适应这些群体所采用的各种细胞模型的不同要求。在 Z 维度中,分辨率应足以解析每个单元的多个平面,而在 X/Y 维度中,需要高分辨率但不一定是超分辨率。总体而言,这些特性应在空间分辨率和时间分辨率之间提供平衡。应可实现超过 10 Hz 的帧速率(例如,记录胞质钙的变化以及线粒体或分泌颗粒的移动性),而不需要导致光漂白或光毒性的荧光激发强度。避免毒性也与连续灌注细胞或组织的长期测量相关。这些功能应该具有足以处理 3D 模型器官(类器官)的穿透深度。转盘共焦显微镜可以满足这些要求。与一次顺序激发一个图像点的荧光的共焦激光扫描显微镜的“传统”变体相比,旋转盘(Nipkow盘)变体凭借包含螺旋形针孔图案的旋转盘一次激发多个荧光图像点。图像生成从一开始就是二维的,允许使用相机芯片作为检测元件。最初,旋转盘原理受到透光率低的困扰。两项发展使旋转盘成为高灵敏度荧光检测仪器。首先,激发光通过平行第二盘中相应的微透镜图案聚焦在针孔图案上。其次,sCMOS 相机的最新发展大大提高了检测效率,该相机结合了非常高的灵敏度和较小的像素尺寸。由于在图像生成过程中物平面中的每个点都会被多次扫描(取决于圆盘的旋转速度),因此每个图像点和时间的激发能量比传统激光扫描显微镜低得多。这使得能够在没有光毒性和/或高图像采集率的情况下进行长期观察。后一个功能可用于细胞内的快速 z 堆叠,从而能够描述 3D 样本中的功能变化而不会出现锯齿。总而言之,旋转盘原理在其当前的发展状态下已经产生了一种非常通用的方法来描述具有高空间和时间分辨率的功能变化。
项目成果
期刊论文数量(0)
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其他文献
吉治仁志 他: "トランスジェニックマウスによるTIMP-1の線維化促進機序"最新医学. 55. 1781-1787 (2000)
Hitoshi Yoshiji 等:“转基因小鼠中 TIMP-1 的促纤维化机制”现代医学 55. 1781-1787 (2000)。
- DOI:
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LiDAR Implementations for Autonomous Vehicle Applications
- DOI:
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2021 - 期刊:
- 影响因子:0
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吉治仁志 他: "イラスト医学&サイエンスシリーズ血管の分子医学"羊土社(渋谷正史編). 125 (2000)
Hitoshi Yoshiji 等人:“血管医学与科学系列分子医学图解”Yodosha(涉谷正志编辑)125(2000)。
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Effect of manidipine hydrochloride,a calcium antagonist,on isoproterenol-induced left ventricular hypertrophy: "Yoshiyama,M.,Takeuchi,K.,Kim,S.,Hanatani,A.,Omura,T.,Toda,I.,Akioka,K.,Teragaki,M.,Iwao,H.and Yoshikawa,J." Jpn Circ J. 62(1). 47-52 (1998)
钙拮抗剂盐酸马尼地平对异丙肾上腺素引起的左心室肥厚的影响:“Yoshiyama,M.,Takeuchi,K.,Kim,S.,Hanatani,A.,Omura,T.,Toda,I.,Akioka,
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