Exciplex detection: application of (i) novel detector systems and (ii) software for signal extraction from noise

Exciplex 检测：应用 (i) 新型检测器系统和 (ii) 从噪声中提取信号的软件

• 批准号: BB/E000223/1
• 负责人: Kenneth Thomas Douglas
• 金额: $ 11.01万
• 依托单位: University of Manchester
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2006
• 资助国家: 英国
• 起止时间: 2006 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FE000223%2F1
• 关键词: Exciplex; detection; application; novel; detector

One of the major scientific advances of recent years has been the inreasing use of information based on DNA samples to help in decision-making in many aspects of everyday life. Immediately obvious examples are the forensic uses of DNA, or medical uses, such as methods for diagnosing disease or potential pathogens. The determination of DNA sequence of the human genome recently has been followed by many similar genome deteminations that should serve to improve or health and safety. A huge number of methods to detect particular regions of a DNA sample, such as from a patient or a potential disease-causing organism, depend on our ability to detect light emission called fluorescence. Naturally, the aim is use as little material as possible in any detemination and for this reason we are seeking ways to minimise any background fluorescence that the DNA analysis method possesses. At Manchester University a new method has recently been developed in which two molecules that do not have any intrinsic fluorescence are brought together on the particular sequence of DNA that is to be detected. The detection molecules have to be very precisely arranged in space for successful fluorescence emission. This correct arrangement of the detector molecules is actually enforced by the DNA target sequence itself. The background fluorescence in this system is less than 1% (this can be compared with other current fluorescence probes for DNA that typically have backgrounds of greater than 60%). By careful design of the chemical structures of these two probe molecules, the system is only able to emit strong fluorescence when exactly the correct DNA sequence has been found. If even a single DNA base is incorrect in the sample sequence, the fluorescence emission is not detectable. This new project extends the scope of these target-assembled exciplex detection of DNA sequences using input of Leicester Space Centre scientists and Edinburgh University astrophysicists. Since 2001 the University of Leicester Space Research Centre (SRC) and Department of Biology, together with the Space Science Department at ESA/ESTEC, have been investigating the application of detectors developed for space astronomy to optical fluorescence measurements in the life sciences and medicine. Their work with superconducting tunnel junctions (STJs) has led to an STJ-based 'scanner' to replace the current types of detectors (CCD- and photomultiplier tube (PMT)-based) for the readout of microarrays or gene chips, for cellular imaging, protein arrays, flow cytometry and many other applications. The STJ offers sensitivity advantages of at least 100 times compared to a silicon CCD and conventional photomultiplier tubes (PMTs) while uniquely measuring the spectral form of fluorescence intensity on a photon-by-photon basis. This new system wil be applied to the DNA exciplexes and allow not only more sensitive measurment, but also a detection system that provides different information from instruments previously used in this context. The above detection methods have concerned themselves with the intensity of the colour of the fluorescence light. However, fluorescence has an additional property - it takes place over a very short (nanosecond), but discrete and measurable, time period. The pattern of the time dependence for these DNA exciplexes is rather complex and this very complexity allows its potential use as a unique badge of the presence or otherwise of such an exciplex in an unknown sample. The detemination of unique patterns of timed events is a common problem in cosmology and so the astronomy groups at Edinburgh University are going to apply their specialist mathematical methods to developing new ways to detect the exciplex time signarure. It is likely that detection using this novel approach will allow the system to be used for very low concentratins - possibly lower than could ever be used based on fluorescence colour insensity alone.

近年来的重大科学进步之一是越来越多地使用基于 DNA 样本的信息来帮助日常生活许多方面的决策。最明显的例子是 DNA 的法医用途或医疗用途，例如诊断疾病或潜在病原体的方法。最近，人类基因组 DNA 序列的测定出现了许多类似的基因组测定，这些测定应该有助于改善健康和安全。检测 DNA 样本特定区域（例如来自患者或潜在致病生物体）的大量方法取决于我们检测称为荧光的光发射的能力。当然，我们的目标是在任何测定中使用尽可能少的材料，因此我们正在寻找方法来最大限度地减少 DNA 分析方法所具有的任何背景荧光。曼彻斯特大学最近开发了一种新方法，将两个不具有任何内在荧光的分子聚集在待检测的特定 DNA 序列上。检测分子必须非常精确地排列在空间中才能成功发射荧光。检测器分子的这种正确排列实际上是由 DNA 目标序列本身强制执行的。该系统中的背景荧光小于 1%（这可以与背景通常大于 60% 的其他当前 DNA 荧光探针进行比较）。通过仔细设计这两种探针分子的化学结构，系统只有在准确找到正确的 DNA 序列时才能发出强烈的荧光。即使样品序列中有一个 DNA 碱基不正确，也无法检测到荧光发射。这个新项目利用莱斯特航天中心科学家和爱丁堡大学天体物理学家的输入，扩展了这些目标组装激基复合物检测 DNA 序列的范围。自 2001 年以来，莱斯特大学空间研究中心 (SRC) 和生物系与 ESA/ESTEC 空间科学系一起，一直在研究为空间天文学开发的探测器在生命科学和医学中光学荧光测量中的应用。他们在超导隧道结 (STJ) 方面的工作催生了基于 STJ 的“扫描仪”，以取代当前类型的探测器（基于 CCD 和光电倍增管 (PMT) 的），用于读取微阵列或基因芯片，用于细胞成像、蛋白质阵列、流式细胞术和许多其他应用。与硅 CCD 和传统光电倍增管 (PMT) 相比，STJ 的灵敏度优势至少高 100 倍，同时以逐个光子为基础独特地测量荧光强度的光谱形式。这个新系统将应用于 DNA 激基复合物，不仅可以进行更灵敏的测量，而且还可以提供与以前在此背景下使用的仪器不同的信息的检测系统。上述检测方法涉及荧光颜色的强度。然而，荧光还有一个额外的特性——它发生在非常短（纳秒）但离散且可测量的时间段内。这些 DNA 激基复合物的时间依赖性模式相当复杂，这种复杂性使其有可能用作未知样品中此类激基复合物是否存在的独特标记。定时事件的独特模式的确定是宇宙学中的一个常见问题，因此爱丁堡大学的天文学小组将应用他们的专业数学方法来开发检测激基复合物时间签名的新方法。使用这种新颖方法的检测很可能允许该系统用于非常低的浓度——可能低于单独基于荧光颜色不敏感所使用的浓度。