Friedrich Mike, Nozadze Revaz, de Keijzer Sandra, Steinmeyer Ralf, Ermolayev Vladimir, Harms Gregory S
Molecular Microscopy Group, Rudolf Virchow Center, University of Würzburg, Josef-Schneider-Str. 2, D15, 97080, Würzburg, Germany.
J Fluoresc. 2018 Jan;28(1):29-39. doi: 10.1007/s10895-011-0966-4. Epub 2011 Oct 6.
Single molecule detection and tracking provides at times the only possible method to observe the interactions of low numbers of biomolecules, inlcuding DNA, receptors and signal mediating proteins in living systems. However, most existing imaging methods do not enable both high sensitivity and non-invasive imaging of large specimens. In this study we report a new setup for selective plane illumination microscopy (SPIM), which enables fast imaging and single molecule tracking with the resolution of confocal microscopy and the optical penetration beyond 300 μm. We detect and report our instrumental figures of merit, control values of fluorescence properties of single nano crystals in comparison to both standard widefield configurations, and also values of nanocrystals in multicellular "fruiting bodies" of Dictyostelium, an excellent control as a model developmental system. In the Dictyostelium , we also report some of our first tracking of single nanocrystals with SPIM. The new SPIM setup represents a new technique, which enables fast single molecule imaging and tracking in living systems.
单分子检测与追踪有时提供了观察活体系统中少量生物分子(包括DNA、受体和信号介导蛋白)相互作用的唯一可行方法。然而,大多数现有的成像方法无法同时实现对大型标本的高灵敏度和非侵入性成像。在本研究中,我们报告了一种用于选择性平面照明显微镜(SPIM)的新装置,它能够以共聚焦显微镜的分辨率进行快速成像和单分子追踪,并且光学穿透深度超过300μm。我们检测并报告了我们仪器的品质因数,与标准宽场配置相比,给出了单个纳米晶体荧光特性的控制值,以及在作为模型发育系统的极佳对照——盘基网柄菌的多细胞“子实体”中纳米晶体的值。在盘基网柄菌中,我们还报告了我们首次使用SPIM对单个纳米晶体进行的一些追踪。新的SPIM装置代表了一种新技术,它能够在活体系统中进行快速的单分子成像和追踪。