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

项目摘要 近年来，人们越来越担心数据的严密性和可靠性来自于 常见和关键的动物模型实验。事实上，科学的一个核心原则--重现性--已经 之所以受到质疑，很大程度上是因为在复制复杂的生物动物实验方面存在挑战。 我们开发了一种快速、便携、定量的聚合酶链式反应(目前通用的“金标准”方法)。 用于各种诊断设置。我们相信，我们的快速、手持、电池供电和 无线设备有可能在世界各地实现快速诊断。在这项提案中，我们的目标是 使用真实世界的样本验证我们的原型设备用于微生物病原体检测。 关于我们的创新技术，我们采取了一种根本不同的取暖和 冷却被放大的样品，避免了标准的帕尔蒂埃加热块。热传递是这样的 效率高，我们能够在短短15秒(甚至可能是 更快)，导致40个周期的运行在大约11分钟内完成。 从本质上讲，这意味着你可以得到关于你的存在和丰富程度的定量答案 目标病原体的最终敏感性明显快于目前的技术水平。 更深入地研究动物数据可变性的根本原因，指出了几种可能的原因 环境和生物原因。在生物学方面，生活在体内或体外的微生物 已被证明对实验结果有明显和目前的影响，产生的数据不一致 有问题。特别是当感兴趣的研究通过设计是翻译的，因此打算 在有益/改善人类健康的同时，还有更大的理由令人担忧。 我们准备测试我们的设备，以识别和量化病原体的存在(或不存在)在一个完整的 在加州大学伯克利分校运营学术动物设施。我们与实验动物关爱办公室一起， 已经确定了两种重要和易处理的病原体作为初步原则研究的良好候选者： 小鼠体内的螺杆菌(细菌)和树突螺旋体，这是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.