基于微流控技术的单细胞操纵和培养研究

Cells in any clonal population can display profound variations on all levels for a variety reasons. Individual cells may play an important role and influence the proliferation of the whole cell population. Conventional techniques for single-cell analysis usually make bulk measurements of a large number of cells and process the collected data on the population levels with statistical methods, which average out the behavior of individual cells. Moreover, cell samples are, in most cases, measured at defined time points when passing through the instruments, so that the resulting data only reflect momentary cellular information. Therefore, in order to obtain precise and dynamic information of individual cells, single-cell analysis is important in the field of biology, medicine and pharmacology. With the rapid development of microfabrication techniques in the last two decades, the concept called ‘lab-on-a-chip’ or ‘microfluidics’ has been increasingly attracting great interest in the research of single-cell analysis. Microfluidics-based single-cell analysis provides various advantages due to the miniaturized geometry close to cell sizes. Microfluidic devices are capable of microfluidic environmental mimesis for cell culturing, and allow for manipulation and detection of single cells. To study single cells in microfluidic systems, cells from a defined population have to be isolated, captured, and cultivated under precisely controlled conditions. The captured cells have to be monitored during the extended culturing period for analysis. .Herein, we propose a high-throughput and -integrity microfluidic device which enables single-cell manipulation and cultivation. The major novelties of this project consist of: (1) capturing a certain number of single cells in suspension; (2) releasing the immobilized cells selectively by using the microelectrode array; (3) real-time monitoring the immobilized cells by using optical microscope. .The core of this research project is to design, develop and experimentally validate a microfluidic device for single-cell manipulation and cultivation. This project, in detail, consists of: (1) the design of a microfluidic configuration for single-cell capture; (2) the design of an addressable microelectrode array for single-cell release; (3) the experimental validation of single-cell manipulation and cultivation in the microfluidic device; (4) the optimization of the microfluidic device, so that the researchers in biology and medicine can operate this device conveniently..

生物体中少量细胞个体对整个细胞群落的生长繁殖有着深远的影响，例如癌变细胞对生命体的影响，因此单细胞分析对生物学，医学及药学研究有着极其重要的意义。传统细胞分析方法往往只对细胞群落进行检测和统计学分析，无法得到动态的单细胞信息；而先进的微加工技术可以实现器件的微型化和精确控制，还能集成多种功能，为单细胞分析提供了许多优势，因此基于微流控技术的单细胞分析近来成为国际研究的热点。本项目提出了一种具有较高通量和集成度的微流控器件以供单细胞操纵和培养，其主要创新点包括：① 可捕获一定数量的单细胞；② 可选择性释放被捕获的单细胞；③ 可对捕获的细胞进行实时监测。本项目的核心研究内容是设计、开发和实验验证该微流控器件平台，具体包括：① 用于单细胞捕获的微流控结构设计；② 用于单细胞释放的可寻址微电极阵列设计；③ 验证性实验；④ 器件优化及操控系统设计，以便生物学医学研究者可方便使用该平台。

通过本项目的实施，我们设计并制作了一种具有较高通量和集成度的微流控器件，并验证了该器件具备单细胞操纵和培养的功能。其主要特点包括：（1）可捕获96个单细胞；（2）可利用介电泳力释放被捕获的细胞；（3）可对被捕获的细胞进行原位对照培养及实时光学监测。