Institute for Neurobiology, University of Münster, Münster, Germany.
PLoS One. 2013;8(1):e53963. doi: 10.1371/journal.pone.0053963. Epub 2013 Jan 21.
We designed a novel imaging technique based on frustrated total internal reflection (FTIR) to obtain high resolution and high contrast movies. This FTIR-based Imaging Method (FIM) is suitable for a wide range of biological applications and a wide range of organisms. It operates at all wavelengths permitting the in vivo detection of fluorescent proteins. To demonstrate the benefits of FIM, we analyzed large groups of crawling Drosophila larvae. The number of analyzable locomotion tracks was increased by implementing a new software module capable of preserving larval identity during most collision events. This module is integrated in our new tracking program named FIMTrack which subsequently extracts a number of features required for the analysis of complex locomotion phenotypes. FIM enables high throughput screening for even subtle behavioral phenotypes. We tested this newly developed setup by analyzing locomotion deficits caused by the glial knockdown of several genes. Suppression of kinesin heavy chain (khc) or rab30 function led to contraction pattern or head sweeping defects, which escaped in previous analysis. Thus, FIM permits forward genetic screens aimed to unravel the neural basis of behavior.
我们设计了一种基于全内反射的新型成像技术（FTIR），以获得高分辨率和高对比度的电影。这种基于 FTIR 的成像方法（FIM）适用于广泛的生物学应用和广泛的生物体。它在所有允许检测荧光蛋白的波长下工作。为了展示 FIM 的好处，我们分析了大量爬行的果蝇幼虫。通过实现一个新的软件模块，该模块能够在大多数碰撞事件中保留幼虫的身份，可分析的运动轨迹数量增加了。该模块集成在我们的新跟踪程序 FIMTrack 中，该程序随后提取了分析复杂运动表型所需的许多特征。FIM 能够实现高通量筛选，即使是细微的行为表型也能筛选出来。我们通过分析几个基因的神经胶质敲低引起的运动缺陷来测试这个新开发的装置。肌球蛋白重链（khc）或 rab30 功能的抑制导致收缩模式或头部清扫缺陷，这在以前的分析中被遗漏了。因此，FIM 允许进行旨在揭示行为神经基础的正向遗传筛选。
