Upconversion nanoparticles as the basis for versatile intracellular luminescence bioassays

拟议的工作的目的是学习建立多功能纳米粒子（NP）平台的动态过程的细胞内分析，并在细胞过程中的干预。发光NP的独特的光学共振能量转移性质允许在局限于NP表面的距离尺度上局部询问选择性生物分子相互作用，并且发光还可以用于诱导可以释放货物或激活与NP相关的生物探针的光反应。 通过用低功率激光照射使纳米颗粒发光。 朝向红外的激发波长将非常适合于穿透到复杂的生物样品中，并且将减少背景散射和自发荧光，但是通常提供不足以激发电子激发以实现期望的光学询问过程的能量。光学地切换生物探针的功能性或释放缀合至NP的货物的期望也受到在长波长下用于激活光反应的能量不足的限制。将近红外（NIR）激发辐射转换为紫外（UV）和可见光的一种优雅方法可以通过上转换过程来实现，其中两个或更多个光子被吸收然后组合。这发生在含有镧系元素的上转换纳米颗粒（UCNP）内。UCNP可以作为供体通过发光共振能量转移（LRET）将能量转移到邻近受体，提供仅对分子距离处的相互作用敏感的转导方法，并提供驱动UCNP表面上的光裂解反应的机会。研究工作将解决在这种技术有效用于细胞内研究之前必须克服的问题，包括： 项目1核-壳-壳UCNP设计，可以在所需波长下稳定高发光强度，同时管理LRET缀合物的空间接近性。 项目2通过使用与UCNP组装的共振耦合金纳米棒增强与激发相关的电场强度来提高UCNP的发光强度。 项目3用多齿膦酸基配体与聚乙二醇的组合涂层改善UCNP在生物相关溶液中的胶体稳定性。 项目4由UCNPs的紫外线辐射引起的聚合物涂层内的光反应，以释放货物或按需激活生物探针。 长期目标是生产一种用于细胞内应用的多功能工具。这些项目的短期目标是学习如何建造发射率更高的UCNPs，并在这些UCNPs上涂上可以同时执行多项任务的组件。

The intention of the proposed work is to learn to build multi-functional nanoparticle (NP) platforms for intracellular analysis of dynamic processes, and for interventions in cellular processes. The unique optical resonance energy transfer properties of luminescent NPs allow for local interrogation of selective biomolecular interactions at a distance scale confined to the surface of the NP, and the luminescence can also be used for induction of photo-reactions that can release a cargo or activate a bioprobe associated with the NP. NPs are made to luminescence by irradiation with a low-power laser. Excitation wavelengths towards the infrared would be well suited for penetrating into complex biological samples and would reduce background scatter and autofluorescence, but typically offer insufficient energy to stimulate electronic excitation to achieve the desired optical interrogation processes. A desire to optically switch on the functionality of bioprobes or to release a cargo conjugated to NPs also is limited by insufficient energy at long wavelengths for activation of photo-reactions. An elegant approach to convert near-infrared (NIR) excitation radiation into ultraviolet (UV) and visible light can be achieved by an upconversion process in which two or more photons are absorbed and then combined. This happens within upconversion nanoparticles (UCNPs) containing lanthanides. UCNPs can serve as donors to transfer energy to proximal acceptors via luminescence resonance energy transfer (LRET), providing a transduction method that is sensitive only to interactions at molecular distances, and affording opportunity to drive photo-cleavage reactions on the surface of UCNPs. The research work will address issues that must be overcome before such technology will be effective for intracellular studies, including: Project 1 Core-shell-shell UCNP designs that can stabilize high luminescence intensity at desired wavelengths while concurrently managing the spatial proximity of conjugates for LRET. Project 2 Improvement of luminescence intensity from UCNPs by increasing the electric field strength associated with excitation using resonance-coupled gold nanorods assembled with UCNPs. Project 3 Improving colloidal stability of UCNPs in biologically-relevant solutions with coatings of multi-dentate phosphonate-based ligands in combination with polyethylene glycol. Project 4 Photo-reactions within polymeric coatings caused by UV emission from UCNPs to release cargo or activate bioprobes on-demand. The long-term goal is to produce a versatile tool for intracellular applications. The short-term objectives of the projects are to learn how to build UCNPs of higher emissivity and to coat these UCNPs with assemblies that can concurrently perform multiple tasks.