Deep and high-resolution three-dimensional tracking of single particles using nonlinear and multiplexed illumination
Deep and high-resolution three-dimensional tracking of single particles using nonlinear and multiplexed illumination
复制标题
DOI：
10.1038/ncomms8874
复制
发表时间：
2015-07-01
影响因子：
16.6
通讯作者：
Dunn, Andrew K.
中科院分区：
文献类型：
--
作者：
Perillo, Evan P.;Liu, Yen-Liang;Dunn, Andrew K.
细胞、组织和工程化三维多细胞模型内的分子运输对于理解包括癌症在内的各种疾病的发展和治疗至关重要。然而，目前的跟踪方法要么局限于两个维度，要么局限于类似于15 μ m的询问深度。在这里，我们提出了一个三维的跟踪方法，能够量化快速的分子输运动力学在高度散射的环境中，深度可达200 μ m。该系统的响应时间为1ms，在高信噪比条件下的时间分辨率可达50 μ s，空间定位精度可达35 nm。这种方法建立在时空复用双光子激发的基础上，只需要一个探测器来进行三维粒子跟踪，并允许双光子多色成像。在这里，我们展示了三维跟踪表皮生长因子受体复合物在肿瘤球体的深度类似于100 μ m。
Molecular trafficking within cells, tissues and engineered three-dimensional multicellular models is critical to the understanding of the development and treatment of various diseases including cancer. However, current tracking methods are either confined to two dimensions or limited to an interrogation depth of similar to 15 mu m. Here we present a three-dimensional tracking method capable of quantifying rapid molecular transport dynamics in highly scattering environments at depths up to 200 mu m. The system has a response time of 1ms with a temporal resolution down to 50 mu s in high signal-to-noise conditions, and a spatial localization precision as good as 35 nm. Built on spatiotemporally multiplexed two-photon excitation, this approach requires only one detector for three-dimensional particle tracking and allows for two-photon, multicolour imaging. Here we demonstrate three-dimensional tracking of epidermal growth factor receptor complexes at a depth of similar to 100 mu m in tumour spheroids.