Menendez-Miranda Mario, Costa-Fernández José Manuel, Encinar Jorge Ruiz, Parak Wolfgang J, Carrillo-Carrion Carolina
Fachbereich Physik, Philipps Universität Marburg, 35037 Marburg, Germany.
Department of Physical and Analytical Chemistry, University of Oviedo, 33006, Spain.
Analyst. 2016 Feb 21;141(4):1266-72. doi: 10.1039/c5an02405f.
Optical analysis based on fluorescence labeling has been extensively used for the selective tagging of a wide range of biomedical important targets or for sensing purposes. Fluorescent nanoparticles (NPs) offer interesting properties as labels, as they can be also used as active labels that change their properties upon changes in the environment, such as pH- or distance-dependent fluorescence. In case NPs are not intrinsically fluorescent, they can be made fluorescent by attaching fluorophores to their volume and/or surface. Dye-labelled NPs can produce a highly amplified optical signal compared to a single dye molecule, as there are many dye molecule attached to each NP, providing a great improvement in analytical sensitivity. However, an appropriate control to quantify the fluorophore/NP ratio is required to succeed in the preparation of quantitative platforms matching the required application. Here a methodology to determine such parameter, the fluorophore/NP ratio, is presented. The methodology combines data obtained from UV/Vis absorption spectroscopy for determination of the dye concentration and inductively coupled plasma-mass spectrometry (ICP-MS) analysis for determination of the NP concentration. To validate the approach, it has been applied to the analysis of different sets of fluorophore-NP conjugates prepared using diverse fluorescent dyes (i.e. fluorophores with different structures and emissions) and several types of NPs (i.e. PbS QDs, Au NPs and FePt NPs). The fluorophore-NP conjugates hereby were designed to incorporate the dye directly into an amphiphilic polymer coating. The developed methodology allows for quantification of fluorophore-NP coupling, and therefore, opens up the possibility of selecting controlled conjugates.
基于荧光标记的光学分析已被广泛用于多种生物医学重要靶点的选择性标记或传感目的。荧光纳米颗粒（NPs）作为标记物具有有趣的特性，因为它们还可以用作活性标记物，在环境变化（如pH或距离依赖性荧光）时改变其性质。如果纳米颗粒本身不具有荧光性，可以通过在其体积和/或表面附着荧光团使其具有荧光性。与单个染料分子相比，染料标记的纳米颗粒可以产生高度放大的光学信号，因为每个纳米颗粒上附着有许多染料分子，从而大大提高了分析灵敏度。然而，为了成功制备符合所需应用的定量平台，需要进行适当的控制以量化荧光团/纳米颗粒的比例。本文介绍了一种确定该参数（荧光团/纳米颗粒比例）的方法。该方法结合了从紫外/可见吸收光谱获得的数据以测定染料浓度，以及电感耦合等离子体质谱（ICP-MS）分析以测定纳米颗粒浓度。为了验证该方法，已将其应用于分析使用不同荧光染料（即具有不同结构和发射的荧光团）和几种类型的纳米颗粒（即PbS量子点、金纳米颗粒和FePt纳米颗粒）制备的不同组荧光团-纳米颗粒缀合物。此处设计的荧光团-纳米颗粒缀合物是将染料直接掺入两亲性聚合物涂层中。所开发的方法能够对荧光团-纳米颗粒的偶联进行定量，因此，为选择可控缀合物开辟了可能性。
