基于光标测技术的激光诱导纳米金纳观热效应测量方法研究

Upon laser heating, the GNP and the surrounding medium experience a number of different thermophysical responses depending on the laser power and GNP, leading to many biological responses and applications at the nano, micro and macroscale, in which the nanoscale responses and applications are most attractive and one of the major challenge. Based on the research that the cell membrane permeability can be elevated by laser activated GNPs bound on the cell membrane,aimed to overcome the difficulties in direct measurement of the nanoscale response, in this project, non-invasive single cell optical mapping technology will be used to measure the membrane voltage change that relative to the membrane permeability. Then a model is presented to describe the relationship between the membrane voltage and the gold nanoscale heating response in order to measure the nanoscale response. We will integrate the laser irradiation system and optical mapping system together, focus on the real time data collection and analysis of the laser activated membrane voltage, present a model describing the relationship between the membrane voltage and membrane permeability, and derive the interaction of membrane voltage change with gold nanoparticles nanoscale heating response under different the laser parameter irradiation. This indirect approach is easier to realize, which can provide a new idea for other related fields.

在不同功率激光照射下，纳米金与周围介质会经历不同的物理学效应，并产生宏观、微观和纳观三个层次的生物学效应与应用。其中纳观效应的生物学应用和机理研究最吸引人并最具挑战性。本项目立足于激光照射结合于细胞膜上的纳米金可使细胞膜穿孔的研究基础，针对纳观效应难以直接观测和研究的难题，提出利用非侵入细胞级光学标测方法检测细胞膜电位变化来间接反应纳米金纳观热效应，并建立相应模型获得细胞膜电位变化与纳米金纳观热效应之间的关系，实现纳米金激光纳观热效应高时空分辨率测量。通过系统的设计与构建，将细胞激光照射系统和光学标测系统高效整合，重点进行激光照射后细胞膜电位的实时采集与分析，建立三维动态细胞通道模型，结合纳米金激光热效应产生的蛋白质变性及气泡产生模型，进而推导出不同参数激光照射下细胞膜电位变化与纳米金纳观效应之间的相互关系模型。这种便捷的检测方法为研究纳米金纳观热效应提供了一种新思路。

为了研究纳米金在不同功率激光照射下的不同生物物理学效应，项目提出在利用激光照射结合于细胞膜上的纳米金产生细胞膜光穿孔的同时，利用非侵入性细胞级光学标测方法监测细胞膜电位变化，并将两者结合起来建立起膜电位变化与膜穿孔之间的模型。主要工作包括：首先采用种子生长法制备纳米金，并把纳米金与抗体结合，检测其结合有效性，并对其是否内吞进行进一步观测研究。其次，对用于光标测的电压敏感染料进行选择并进行性能测试，搭建双激光荧光显微成像系统，并对激光诱导纳米金致细胞膜穿孔后膜电位的测量。然后进行优化，获得较好的实验结果。最后建立细胞膜电位变化和纳米颗粒与细胞膜作用关系的数学模型，表征其与膜上电位变化的关系。结果表明，光穿孔造成的膜损伤是可恢复的，膜电位的恢复时间与膜穿孔尺寸呈一定的对应关系。在此基础上，本项目还对纳米金光热效应的机理进行了研究，并扩展了其在光动力治疗等方面的应用。