Sub-nm dendrimer-metal nanoclusters as ultrabright, mod*

描述（由申请人提供）：了解生命系统内固有的异质性需要开发新的体内单分子（SM）光学方法来跟踪蛋白质动力学，而不需要集合平均的面纱。贵金属纳米团簇在整个可见光和近红外光谱中表现出极强的尺寸依赖性发射，但其尺寸远小于同类半导体量子点（<l-nm）。金属纳米团簇的高极化率导致极短，高效率的辐射寿命（约30-ps，量子产率约50%），甚至增强了封装支架的拉曼信号，使其在单分子水平上可观察到。我们将继续使用聚（胺胺）树状大分子（PAMAM）来溶解和稳定这些高发射的纳米团簇。通过树状大分子合成，a)在无背景光谱区域整合特定的拉曼活性标签，b)模块化树状大分子功能化，以整合广义树状大分子封装纳米团簇（“纳米点”）生化功能，我们将开发出独特的体内单分子成像材料。独特的光物理特性（极快的辐射寿命，高量子产率，无需大的纳米粒子就能产生拉曼信号）使这些亚纳米纳米团簇与更大的（3-10纳米）半导体量子点一样强吸收，但是，由于它们不受大约10纳米的长量子点辐射寿命的限制，纳米点的发射速率和亮度至少高出两个数量级。我们将充分表征这种新型重要纳米材料的光学响应，因为我们将它们用作生物标签。它们的优势特性使时间和光谱门控检测能够获得非常高的单分子信号，即使在活细胞特征的高自荧光背景存在的情况下。

DESCRIPTION (provided by applicant): Understanding the inherent heterogeneity within living systems demands the development of new in vivo single molecule (SM) optical methods to follow protein dynamics without the veil of ensemble averaging. Noble metal nanoclusters exhibit exceedingly strong, size dependent emission throughout the visible and near IR spectrum, but at much smaller sizes (<l-nm) than comparable semiconductor quantum dots. The high polarizability of metal nanoclusters leads to extremely short, high efficiency radiative lifetimes (approximately 30-ps, and quantum yield of approximately 50%), and even enhances the Raman signal from the encapsulating scaffold to make it observable on the single molecule level. We will continue using poly(amidoamine) dendrimers (PAMAM) to solubilize and stabilize these highly emissive nanoclusters. Through dendrimer synthesis to a) incorporate specific Raman active labels in background-free spectral regions and b) modular dendrimer functionalization for incorporating generalized dendrimer encapsulated nanocluster ('nanodot') biochemical functionality, we will develop materials that uniquely enable in vivo single molecule imaging. The unique photophysics (extremely fast radiative lifetime, high quantum yield, and ability to produce Raman signals without a large nanoparticle) make these sub-nm nanoclusters as strongly absorbing as much larger (3-10 nm) semiconductor quantum dots, but, because they are not limited by the long quantum dot radiative lifetime of approximately 10 ns, the nanodot emission rates, and therefore brightness are at least two orders of magnitude higher. We will fully characterize the optical response of this new class of important nanomaterials as we employ them as biological labels. Their advantageous properties enable time and spectrally gated detection to obtain very high single molecule signals even in the presence of high autofluorescent backgrounds characteristic of living cells. We have assembled an outstanding team to chemically functionalize the PAMAM scaffold encapsulating and stabilizing the highly emissive Au and Ag nanoclusters and optically and biochemically characterize them in vitro and in vivo. Through three specific Aims, we will develop these robust ultrabright and ultrasmall nanodots into unparalled, specific, in vivo biological labels. In Aim I we will use targeted chemical synthesis to incorporate modularity in the PAMAM scaffold such that any functional group for modular attachment of biochemical targeting and recognition units can be incorporated. In Aim II we will synthesize and attach membrane transport functionalities to the nanodots and characterize their uptake and optical properties. These studies lead to Aim III in which multifunctionalized nanodots are made to specifically bind fusion proteins within the cytosol and we gate their transport into specific organelles. The single molecule imaging methods will be developed such that these extremely bright probes can be directly imaged by temporally and spectrally rejecting essentially all background from the more long-lived (ns) autofluorescent species. These combined methods should be capable of increasing current signal/noise ratios by more than three orders of magnitude over current nanoparticle or organic fluorophore based methods. This toolbox of modular, ultrabright, ultrasmall, and short-radiative lifetime nanodots will be generally applicable to a wide range of systems and will be made available to the community through this Exploratory Center.