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Determination of position and trajectory of vesicles containing nanoparticles using centroid calculation For quantitative determination of the intracellular motion of the nanovesicles, 3D image stacks were acquired at 5-minute intervals. Each stack consists of eight slices with a slice thickness of 0.8 μm. To correct for any macroscopic translation errors between acquisition times, each image stack was rigidly registered to the first acquired stack using a 3D phase correlation technique。After registration, connected 3D fluoresced regions from the first stack were segmented using a flood-filling algorithm. By appropriate choices of the threshold arguments, effects caused by fluorescence background noise can be removed. The thresholds are reduced to mitigate photobleaching effects in stacks acquired after the first.This technique is explained in further detail in the online version of this article (Supplement S2). Denoting the centroid of a nanovesicle at acquisition time ti by Ri, the average speed (Si) of the group at each acquisition time was calculated as （公式）and the 3D diffusion coefficient (Di) at time ti as（公式） The position of a nanovesicle when it is registered in more than one Z-plane was determined by centroidcalculation as discussed in the online version of this article (Supplement S2). The position of each nanovesicle was mapped as a function of time using the above method, to exploit the trajectory and other transport properties (speed, diffusion coefficient) of nanostructures within the cell. |
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zhanchuan
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- 专业: 天然药物化学
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sltmac(金币-5): 本版禁止机器翻译，烦请仔细阅读版规，初犯，扣5BB~~~ 2011-09-29 18:31:00
sltmac(金币-5): 本版禁止机器翻译，烦请仔细阅读版规，初犯，扣5BB~~~ 2011-09-29 18:31:00
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测定的位置和轨迹的质心计算含有纳米粒子的囊泡 定量测定的细胞内的nanovesicles议案，获得三维图像栈，每隔5分钟。每个堆栈由八片的层厚0.8微米。要正确采集时间之间的任何宏观的翻译错误，每个图像堆栈是硬性注册使用三维相位相关技术的第一次收购栈。注册后，从第一个堆栈连接的三维荧光地区分割使用洪水填充算法。通过阈值参数的适当选择，荧光背景噪音所造成的影响可以被删除。阈值降低，以减轻漂白在收购协议栈的影响，在本条第（补编S2）的在线版本的进一步详细解释后first.This技术。表示在收购RI时间Ti nanovesicle的重心，在每次采集时间平均车速组（SI）计算（公式）和三维扩散系数（DI）在时间Ti（公式）的位置当它是在一个以上的Z平面注册nanovesicle由centroidcalculation决定在本条第（补编S2）的在线版本的讨论。每个nanovesicle位置映射为一个时间用上述方法的功能，利用的轨迹和其他细胞内的输运性质的纳米结构（速度，扩散系数）。 |
2楼2011-09-29 12:23:57
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