Polymer Dot Nanoparticles for Detection of Single Molecules in Live Cells

描述（由申请人提供）：拟议研究的目的是开发一类称为“聚合物点”的新型荧光纳米颗粒，并使用常规荧光显微镜方法测试纳米颗粒用于检测活细胞中单个分子的可行性。聚合物点纳米颗粒是一种基于荧光π-共轭聚合物的有前途的新型荧光纳米颗粒技术，其与常规荧光染料和纳米颗粒相比提供了亮度的实质性改善。该项目的具体目标包括优化和表征单分子检测的相关品质因数，开发和测试针对细胞内特定生物分子的生物缀合策略，以及在活细胞中演示单个纳米颗粒检测。将采用一系列表征技术，包括荧光光谱、单分子光谱、原子力显微镜、电泳和落射荧光显微镜。预期与常规荧光染料相比，亮度和光稳定性提高100至10，000倍，满足活细胞中基于荧光的单分子检测的要求。还预计将展示针对感兴趣的特定生物分子的简单、灵活的生物缀合方法以及检测活细胞内单个标记生物分子。这种新型荧光纳米颗粒平台的开发将促进未来的研究，包括监测活细胞内单分子的运输和生化事件，也可以为新型生物测定和生物传感器的开发提供基础。

DESCRIPTION (provided by applicant): The objective of the proposed research is to develop a novel class of fluorescent nanoparticles called "Polymer Dots" and test the feasibility of the nanoparticles for detection of single molecules in live cells using conventional fluorescence microscopy methods. Polymer dot nanoparticles are a promising new fluorescent nanoparticle technology based on fluorescent pi-conjugated polymers which provide substantial improvements in brightness as compared to conventional fluorescent dyes and nanoparticles. The specific aims of the project include optimizing and characterizing relevant figures of merit for single molecule detection, development and testing of bioconjugation strategies for targeting specific biomolecules within cells, and demonstration of single nanoparticle detection in living cells. A range of characterization techniques will be employed, including fluorescence spectroscopy, single molecule spectroscopy, atomic force microscopy, electrophoresis, and epifluorescence microscopy. Improvements in brightness and photostability of a factor of 100 to 10,000 as compared to conventional fluorescent dyes are expected, meeting the requirements for fluorescence-based single molecule detection in living cells. Demonstration of facile, flexible bioconjugation methods for targeting specific biomolecules of interest and detection of individual labeled biomolecules within living cells are also expected. The development of this novel fluorescent nanoparticle platform will facilitate future research involving monitoring transport and biochemical events of single molecules within living cells and could also provide the basis for the development of novel bioassays and biosensors.