Novel System for Automation of Zebrafish Lipid Assays

描述(申请人提供)：斑马鱼，Danio rerio，已被广泛接受为脊椎动物发育的有用模型。斑马鱼作为一种用于高通量药物发现分析的模式生物引起了人们的强烈兴趣。到目前为止，斑马鱼真正的高通量筛选仅限于胚胎和非常早期的幼体。这是因为研究人员最感兴趣的许多器官最好是从动物的角度来观察。一旦鱼囊充气，幼虫通常直立漂浮，就很难产生动物侧面的清晰图像。在多孔微板格式中尤其如此。解决这一重要问题的努力集中在使用共焦成像技术或强大的图像去卷积软件来组装动物的三维重建。这些努力没有达到所需的空间分辨率，通常需要1到2分钟才能成像一口井。物理科学公司(PSI)发明了一种新技术，该技术将适用于已经在药物发现实验室使用的标准图像分析系统。PSI的技术将允许研究人员在微孔阵列中生成年龄较大的斑马鱼幼体的清晰侧视图。这项技术不需要共焦显微镜技术或复杂的图像去卷积算法，因此每井几秒钟就能产生高质量的图像，而其他技术每井几分钟。在第一阶段，我们使用了Zygogen公司正在开发的Z-LipoTrack分析来证明我们技术的可行性。之所以选择脂迹分析，是因为它通常是在较大的幼虫身上进行的，需要收集胆汁和肠道的图像。在第二阶段，我们将进一步开发PSI的创新成像技术，并将该技术整合到一个完全集成的斑马鱼幼体自动发现系统中。PSI的技术将使用Z-LipoTrack分析实现高通量发现，以降低胆固醇和血脂水平的生物活性化合物。这些药物满足了一个重要的医疗保健需求。与公众健康相关：这项研究将开发能够在活性药物化合物的高通量筛选中使用人类疾病的斑马鱼模型的技术。正在开发的这种特殊检测方法有望加速发现降胆固醇药物。我们预计，利用成像技术的分析还将加速发现治疗多种人类疾病的新药物，如心血管疾病和急性肾功能衰竭。

DESCRIPTION (provided by applicant): The zebrafish, Danio rerio, has become widely accepted as a useful model for vertebrate development. There is strong interest in developing zebrafish as a model organism for use in high throughput drug discovery assays. Thus far, true high throughput screening in zebrafish has been limited to embryos and very early larvae. This is because many of the organs that are most interesting to researchers are best viewed from the side of the animal. Once the swim bladder inflates and the larvae generally float upright it is difficult to generate a clear image of the side of the animal. This is especially true in multiwell microplate format. Efforts to resolve this important problem have centered on use of confocal imaging techniques or powerful image deconvolution software to assemble 3-D reconstructions of the animal. These efforts have not achieved the desired spatial resolution and typically require 1 to 2 minutes to image a single well. Physical Sciences Inc. (PSI) has invented a novel technology that will be adaptable to standard image analysis systems that are already in use in drug discovery laboratories. PSI's technology will allow researchers to generate clear side-views of older zebrafish larvae in microwell arrays. The technology does not require confocal microscopy techniques or sophisticated image deconvolution algorithms and will therefore generate quality images in seconds per well versus minutes per well for those other techniques. In Phase I we used the Z-Lipotrack assay under development at Zygogen Corp to demonstrate the feasibility of our technology. The Lipotrack assay was chosen because it is typically performed on older larvae and requires collection of images of the gall bladder and intestine. In Phase II we will further develop PSI's innovative imaging technology and integrate that technology into a fully integrated, automated discovery system for zebrafish larvae. PSI's technology will enable high throughput discovery using the Z-Lipotrack assay of bioactive compounds that reduce cholesterol and lipid levels. Those drugs meet an important health care need. Relevance to public health: This research will develop technologies that will enable the use of zebrafish models of human disease in high-throughput screens for active pharmaceutical compounds. The specific assay under development is expected to accelerate discovery of cholesterol-lowering drugs. We anticipate that assays utilizing the imaging technology will also accelerate discovery of new Pharmaceuticals to treat a broad range of human illness such as cardiovascular disease and acute renal failure.