A portable quantitative polymerase chain reaction platform (qPCR) for rapid detection of pathogens impacting model organisms in animal facilities

项目概要 近年来，人们越来越关注来自数据的严谨性和可靠性。 常见和关键的动物模型实验。事实上，科学的一个核心原则——可重复性——已经 受到质疑主要是由于复制复杂的生物学动物实验面临的挑战。 我们开发了一种快速、便携式、定量 PCR（当前通用的“金标准”方法）设备 适用于各种诊断环境。我们相信我们的快速、手持、电池供电和 无线设备有潜力在世界各地实现快速诊断。在本提案中，我们的目标是 使用真实世界的微生物病原体检测样本来验证我们的原型设备的使用。 关于我们的创新技术，我们采取了根本不同的方法来加热和 冷却正在放大的样品，避免使用标准珀耳帖加热块。传热是这样的 高效，我们能够在短短 15 秒内执行加热和冷却循环（甚至可能 更快），从而在大约 11 分钟内完成 40 个周期的运行。 从本质上讲，这意味着您可以获得有关您的物质的存在和丰富度的定量答案 以比当前最先进技术快得多的最终灵敏度靶向病原体。 深入研究动物源数据变异性的根本原因，指出了几种可能的原因 环境和生物原因。从生物学角度来说，微生物是生活在体内或体外的 事实证明对实验结果有明显而现实的影响，产生的数据不一致且 有疑问。特别是，当感兴趣的研究是有意转化的，因此旨在 有益/改善人类健康，更值得关注。 我们准备测试我们的设备，以全面识别和量化病原体的存在（或不存在）。 在加州大学伯克利分校运营学术动物设施。我们与实验动物护理办公室一起 已经确定了两种重要且易于处理的病原体作为初步原理研究证明的良好候选者： 小鼠体内的螺杆菌（细菌）和树突芽孢杆菌（一种影响两栖动物的真菌）。

Project Summary In recent years there have been growing concerns about the rigor and reliability of data originating from common and critical animal model experimentation. In fact, a core principal of science – reproducibility – has been called into question largely due to challenges in replicating biologically complicated animal experiments. We have developed a fast, portable, quantitative PCR (the current universal “gold standard” method) device for use in a wide variety of diagnostic settings. We believe that our rapid, handheld, battery-powered and wireless device has the potential to empower rapid diagnostics around the world. In this proposal we aim to validate the use of our prototype device using real-world samples for microbial pathogen detection. Regarding our innovative technology, we have taken a fundamentally different approach to heating and cooling the sample being amplified, avoiding the standard Peltier heating block. The heat transfer is so efficient that we are able to perform a heating and cooling cycle in as little as 15 seconds (possibly even faster), resulting in a 40-cycle run finishing in about 11 minutes. In essence, this means you can get a quantitative answer regarding the presence and abundance of your target pathogen with ultimate sensitivity significantly faster than the current state of the art. Looking deeper into the root causes of animal-derived data variability is pointing to several possible environmental and biological causes. In terms of the biology, microbes that live inside or outside of the body are proving to have clear and present effects on experimental outcomes, yielding data that is inconsistent and questionable. In particular, when the studies of interest are translational by design, and therefore intended to benefit / improve human health, there is even greater cause for concern. We are poised to test our device to identify and quantify the presence (or absence) of pathogens in an fully operating academic animal facility at UC Berkeley. Together with the Office of Laboratory Animal Care, we have identified two important and tractable pathogens as good candidates for initial proof of principle studies: Helicobacter species (bacteria) in mice and B. dendrobatidis, a fungus affecting amphibians.