Quantitative Biosensing at the Single-Molecule Level

化学系的化学测量和成像(CMI)项目正在支持犹他大学的Joel M.Harris教授和他的学生开发新的显微镜方法，以帮助理解和优化传感器，以超灵敏地检测目标生物分子，包括DNA和蛋白质。这些目标正在通过单个分子的定量成像来实现。单分子荧光成像可以提供生物传感器表面结合位点和结合靶分子的绝对覆盖信息。这项技术代表了对单个分子进行极限计数的定量分析。它首次被应用于研究与细胞信号和蛋白质在生物传感器表面的行为相关的与脂双层结合的多肽。定量单分子成像也被用于测量DNA杂交，这构成了基因测试和基因芯片应用的基础。人们正在研究静电相互作用对DNA结合的影响，以了解离子强度如何在DNA传感器应用中控制变异性。单分子成像也可以作为一种分析方法，通过计算捕获表面上结合的单个DNA分子的数量，可以检测到生物样本中极低浓度的目标DNA。该计划正在组装能够对单分子成像的新仪器，以及对图像中分子进行定量检测的数据分析方法。这项研究将为人们对影响用于检测疾病和毒素的生物传感器性能的结构性因素提供新的理解。这项研究还促进了对蛋白质-膜相互作用和DNA杂交动力学的基本理解，这对开发新的药物和生物技术至关重要。该研究为研究生和本科生提供光谱学、显微镜和生物分析化学方面的多学科培训，并支持与其他研究小组的几个合作。该项目还通过开发显微镜练习并将其部署在盐湖城一所第一小学的教室中，影响了K-12科学教育和公众意识。

The Chemical Measurement and Imaging (CMI) program of the Division of Chemistry is supporting Professor Joel M. Harris and his students at the University of Utah to develop new microscopy methods to help understand and optimize sensors for ultrasensitive detection of target biomolecules, including DNA and proteins. These goals are being addressed through the quantitative imaging of individual molecules. Single-molecule fluorescence imaging can provide absolute coverage information of both binding sites and bound target molecules on a biosensor surface. This technique represents quantitative analysis at the ultimate limit of counting individual molecules. It is first being applied to studies of peptide binding to lipid bilayers that is relevant to cell signaling and to the behavior of proteins at biosensor surfaces. Quantitative single-molecule imaging is also being adapted to measuring DNA hybridization, which forms the basis of genetic testing and gene-chip applications. The influence of electrostatic interactions on DNA binding is being studied in order to understand how ionic strength acts at controlling variability in DNA sensor applications. Single-molecule imaging can also serve as an analytical method, where counting individual bound DNA molecules on a capture surface enables detection of extremely low concentrations of target DNA in biological specimens. The program is assembling new instrumentation capable of imaging single molecules and data analysis methods for quantitative detection of molecules in the images. This research will provide routes to new understanding of structural factors that influence the performance of biosensors that are used to detect disease and toxins. The research also advances fundamental understanding of protein-membrane interactions and DNA hybridization kinetics, which are critical to development of new pharmaceuticals and biotechnologies. The research provides multidisciplinary training of graduate and undergraduate students in spectroscopy, microscopy, and bioanalytical chemistry and supports several collaborations with other research groups. The project also impacts K-12 science education and public awareness through development of microscopy exercises and their deployment in classrooms of a Title I elementary school in Salt Lake City.