Multiplicative Amplification with Singlet Oxygen and Conjugated Polymers for Bioanalytical Applications

在化学系化学测量和成像（CMI）项目的支持下，塔夫茨大学的Samuel托马斯教授及其团队将开发一种新的荧光检测信号放大方法。 荧光传感器和测定是分析科学中的一组关键技术，并且对特定分子或其他刺激的荧光响应的放大对于许多技术（例如ELISA）所需的高灵敏度是至关重要的。 然而，需要开发新的方法，通过提高定量准确性、在具有挑战性的环境中的鲁棒性以及减少假阳性和假阴性来提高性能。 该项目的目的是测试以下假设：结合两种形式的放大：1）通过光敏化的单线态氧（1 O2）的化学放大，以及2）光捕获和激子迁移率将产生倍增放大的荧光响应，这在生物分析应用中是有用的。 该小组将追求以下两个目标来测试他们的中心假设：1）证明用与单线态氧反应的陷阱取代的共轭聚合物的荧光响应的倍增放大，以及2）使用单线态氧响应聚合物使用夹心测定以高选择性检测1.0 pM或更低的目标生物分子。 这种方法的成功实现将产生独特且有用的特征组合，例如比率响应和不需要大的酶标记，这克服了当前方法的局限性，并且在一系列分析应用中可能有用。 所提出的工作的更广泛的影响将是双重的：1）用于放大荧光信号的新方法具有改进依赖于夹心测定的实验技术的能力，例如临床样品分析和高通量药物发现; 2）一个将有机光化学研究与掩体山社区学院的演示和实验相结合的项目，托马斯教授已经与之有工作关系。信号放大？将少量样本转换为相对较大的读数的过程？是现代样品分析的高灵敏度的潜在关键，无论是在临床还是在现场，在诸如通过荧光分析DNA或蛋白质的应用中。 然而，目前的金标准扩增方法的局限性阻碍了下一代技术的发展，这些技术可以在较低水平上检测靶分子，减少假阳性和假阴性，特别是在缺乏对环境条件控制的诊所之外。 通过将两种已知的信号放大方法与专门设计的荧光材料相结合，这项跨学科的研究将产生一种新的方法来放大荧光信号，这种方法通常可用于一系列生物分析应用。 这种方法相对于现有技术的优点包括1）荧光信号的更稳健的读出方法，2）不需要大的酶标记物来实现扩增，大的酶标记物可能导致测定的稳定性和灵敏度的问题，以及3）由于两种形式的扩增的组合而增加的灵敏度。 除了这些研究成果将为社会带来的好处外，该小组还将把他们的研究纳入波士顿附近社区学院掩体希尔CC的外展计划，该学院80%的学生属于少数群体，超过50%是女性，以开发实验和演示，让学生亲身体验光化学，包括使用他们在研究中开发的材料。

With support from the Chemical Measurement and Imaging (CMI) Program in the Division of Chemistry, Professor Samuel Thomas at Tufts University and his group will develop a new approach to signal amplification in fluorescent assays. Fluorescent sensors and assays are a key set of technologies in analytical science, and amplification of fluorescent response to a specific molecule or other stimulus is critical to the required high sensitivity of many technologies such as ELISA. There is a need, however, to develop new approaches that improve performance through increased quantitative accuracy, robustness in challenging environments, and reduced false positives and negatives. The objective of this project is to test the hypothesis that combining two forms of amplification: 1) chemical amplification of singlet oxygen (1O2) through photosensitization, and 2) light harvesting and exciton mobility will yield multiplicatively amplified fluorescent response that is useful in bioanalytical applications. The group will pursue the following two objectives to test their central hypothesis: 1) Demonstrate multiplicative amplification of fluorescence response of conjugated polymers substituted with traps that react with singlet oxygen, and 2) Use singlet oxygen-responsive polymers to detect target biomolecules at 1.0 pM or lower with high selectivity using sandwich assays. Successful realization of this approach would yield a unique and useful combination of features, such as a ratiometric response and the lack of a requirement for a large enzyme label, which overcomes limitations of current approaches and is potentially useful across a range of analytical applications. The broader impacts of the proposed work will be twofold: 1) a new method for amplifying fluorescent signal has the capability to improve to experimental techniques that rely on sandwich assays such as analysis of clinical samples and high-throughput drug discovery; 2) a program that integrates this research in organic photochemistry with demonstrations and experiments at Bunker Hill Community College, with which Prof Thomas already has a working relationship.Signal amplification?processes that convert a small quantity of sample into a comparatively large readout?are an underlying key to the high sensitivity of modern sample analysis both in the clinic and in the field, in applications such as analysis of DNA or proteins by fluorescence. Limitations of current gold standard methods of amplification, however, are preventing the development of next-generation technologies that can detect targeted molecules at lower levels with reduced false positives and false negatives, especially outside the clinic, where there is a lack of control over environmental conditions. By combining two known methods of signal amplification with specially designed fluorescent materials, this proposed interdisciplinary research will yield a new approach to amplifying fluorescent signal that will be generally useful in a range of bioanalytical applications. Advantages of this approach over state-of-the-art technologies include 1) a more robust readout method of fluorescent signal, 2) no requirement for large enzyme labels, which can cause problems with both stability and sensitivity of an assay, to achieve amplification, and 3) increased sensitivity due to the combination of two forms of amplification. In addition to the benefit to society that such research achievements would provide, the group will also integrate their research into an outreach program at Bunker Hill CC, a nearby community college in Boston where 80% of the students belong to minority groups and over 50% are women, to develop experiments and demonstrations to give students hands-on experience with photochemistry, including using materials they develop in the research.