Novel System for Automation of Zebrafish Lipid Assays

描述（由申请人提供）：斑马鱼（Danio rerio）已被广泛接受为脊椎动物发育的有用模型。人们对开发斑马鱼作为用于高通量药物发现测定的模式生物有强烈的兴趣。到目前为止，真正的高通量筛选斑马鱼已限于胚胎和非常早期的幼虫。这是因为研究人员最感兴趣的许多器官最好从动物的侧面观察。一旦鱼鳔膨胀，幼虫通常直立漂浮，就很难生成动物侧面的清晰图像。在多孔微孔板形式中尤其如此。解决这一重要问题的努力集中在使用共聚焦成像技术或强大的图像去卷积软件来组装动物的3-D重建。这些努力没有实现期望的空间分辨率，并且通常需要1至2分钟来对单个井进行成像。物理科学公司(PSI)发明了一种新技术，将适用于已在药物发现实验室使用的标准图像分析系统。PSI的技术将使研究人员能够在微孔阵列中生成老年斑马鱼幼虫的清晰侧视图。该技术不需要共聚焦显微镜技术或复杂的图像去卷积算法，因此将在每孔数秒内生成高质量的图像，而其他技术则需要每孔数分钟。在第一阶段，我们使用Zygogen公司正在开发的Z-Lipotrack检测来证明我们技术的可行性。选择Lipotrack测定是因为其通常在较老的幼虫上进行，并且需要收集胆囊和肠的图像。在第二阶段，我们将进一步开发PSI的创新成像技术，并将该技术集成到一个完全集成的斑马鱼幼虫自动发现系统中。PSI的技术将使高通量发现使用Z-Lipotrack测定的生物活性化合物，降低胆固醇和脂质水平。这些药物满足了重要的医疗保健需求。与公共卫生的相关性：这项研究将开发技术，使人类疾病的斑马鱼模型能够用于活性药物化合物的高通量筛选。正在开发的特定检测方法有望加速降胆固醇药物的发现。我们预计，利用成像技术的检测也将加速新药的发现，以治疗广泛的人类疾病，如心血管疾病和急性肾衰竭。

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.