A method for single molecule tracking using a conventional single-focus confocal setup
A method for single molecule tracking using a conventional single-focus confocal setup
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DOI：
10.1063/1.5083869
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发表时间：
2019-03-21
影响因子：
4.4
通讯作者：
Presse, Steve
中科院分区：
文献类型：
--
作者：
Jazani, Sina;Sgouralis, Ioannis;Presse, Steve
利用荧光成像实现空间分辨率的一种方法是使用宽视场照明，并通过多个传感器（相机像素）收集测量数据。在这里，我们提出了另一种使用共聚焦测量和单个传感器的方法。传统上，共聚焦显微镜已被用来实现高时间分辨率在空间分辨率的代价。这是因为它使用很少的，通常只有一个传感器来收集数据。然而，共聚焦数据编码空间信息。在这里，我们表明，在形状的共焦激发体积的不均匀性可以利用来实现空间分辨率。为了实现这一目标，我们制定了一个专门的隐马尔可夫模型，并采用前向滤波-后向采样马尔可夫链蒙特卡罗方案，以有效地处理荧光相关光谱中常用的对称共聚焦体积内的分子运动。我们的方法可以用于单共焦体积应用，也可以结合到更大的计算方案中，用于专门的，多共焦体积，光学设置。由AIP出版社授权出版。
One way to achieve spatial resolution using fluorescence imaging-and track single molecules-is to use wide-field illumination and collect measurements over multiple sensors (camera pixels). Here we propose another way that uses confocal measurements and a single sensor. Traditionally, confocal microscopy has been used to achieve high temporal resolution at the expense of spatial resolution. This is because it utilizes very few, and commonly just one, sensors to collect data. Yet confocal data encode spatial information. Here we show that nonuniformities in the shape of the confocal excitation volume can be exploited to achieve spatial resolution. To achieve this, we formulate a specialized hidden Markov model and adapt a forward filtering-backward sampling Markov chain Monte Carlo scheme to efficiently handle molecular motion within a symmetric confocal volume characteristically used in fluorescence correlation spectroscopy. Our method can be used for single confocal volume applications or incorporated into larger computational schemes for specialized, multi-confocal volume, optical setups. Published under license by AIP Publishing.