【作者】 张弦；

【导师】 杜伟；

【作者基本信息】 华中科技大学 ， 生物化学与分子生物学， 2017， 硕士

【摘要】 循环肿瘤细胞(circulating tumor cells,CTCs)作为癌症诊断、预后和疾病监控的重要指示物。外周血中CTCs极度稀少,因此其有效分离非常具有挑战性。当前有两种方法用来分离CTCs,一种是根据细胞大小分离,一种是基于免疫亲和性分离。然而根据当前的捕获效率来看,这些捕获方法都需要改进。本文基于微流控芯片平台,设计一种以仙人掌为原型的高光学透明性的微柱矩阵芯片装置,可从X、Y、Z三个方向高效捕获肿瘤细胞。通过软光刻和快速原型法制作微柱矩阵芯片,同时通过静电化学沉积法在芯片上镀上一层金薄层,使得该微柱表面的金层和微通道具有层次结构。通过含有SH基团的寡核苷酸链将EpCAM抗体连接到微柱和微通道表面带有的微纳尺度金颗粒上,因此CTCs可以同时被微柱和微通道上的EpCAM抗体结合,从而实现CTCs三维捕获。研究表明该装置可以高效捕获不同癌细胞和释放肿瘤细胞,在PBS缓冲液中平均捕获效率高达88%,在全血样本中可达83.7%。另外,整个装置具有高透明度,可在显微镜下监控整个捕获过程,表明该项研究在临床诊断中具有很高的临床价值和应用潜力。本文将温敏性水凝胶和微流控微柱芯片相结合,研究了智能水凝胶的温敏特性在细胞捕获中的应用。将多巴胺加入温敏性水凝胶从而形成具有光热转化性能的复合水凝胶,结果表明加入多巴胺的复合水凝胶的微观结构发生改变,并具有光热性能。同一强度的激光照射170s后,随着多巴胺浓度的上升,复合水凝胶的温度上升也随之更加明显,最高升温可达到30oC;而对照组水凝胶温度几乎没有变动。同时还将该复合水凝胶应用于细胞学研究,如CTCs捕获,肿瘤细胞粘附,细胞密度片层以及特定图案化建立。细胞实验中细胞捕获率达到约84%,粘附效果非常理想,密度片层以及图案片层能够快速且规整的形成,这些显示出该复合水凝胶具有较大的应用潜力。更多还原

【Abstract】 The circulating tumor cell(CTC)is an important indicator in cancer diagnosis,prognosis,and disease monitoring.However,efficient isolation of CTCs from the peripheral bloodstream has been a persistent challenge due to the rarity of CTCs.Currently,two types of microfluidic methods have been reported for the isolation of CTCs,including size-based and immunoaffinity-based microfluidics.However,the capture efficiency of current methods still needs improvement.In this work,we reported a “ Rhipsalis(Cactaceae)”-like micropillar array-based device with high optical transparency for efficiently capturing cancer cells in three dimensions(XY and Z planes).The PDMS micropillar array was fabricated by soft-lithography and rapid prototyping method.The micropillar array was plated with a thin gold layer through electroless plating.EpCAM antibody was then modified onto the surface of the micropillars through the thiol-oligonucleotide linkers.Hierarchical structures originating from the gold layer are on the surface of the micropillars and the microchannel.The immune-affinity molecules(EpCAM antibodies)could be modified on the gold-coated micropillars and microchannel.Thus,capture of cancer cells could be realized simultaneously on the surface of the micropillars and the microchannel.The device in this study demonstrated a high capture efficiency and release for different kinds of cancer cells,the average capture efficiency of 88% was achieved in PBS and the cancer-cell capture capability with average efficiency of 83.7% in whole blood samples.The high transparency of the whole device allows for direct monitoring captured cells under microscopy,showing its potential applications in CTC studies and clinical diagnosis.Based on the thermo-responsive hydrogels and the combination of microfluidic chip platform,we studied the thermo-responsive hydrogels after joining dopamine,the composite hydrogel had the characteristics of photothermal conversion efficiencies and the change of microstructure.Also,we take advantage of the hydrogel’s thermo-responsive characteristics to implement a series experiment about cells,such as capturing cancer tumor cells,adhesioning cancer cells,density slice layer test of cancer cells and the specific pattern layer test of cancer cells.In this study demonstrated that the composite hydrogel had the characteristics of photothermal conversion efficiencies after joining dopamine,when the same intensity of laser irradiate 170 seconds,with the rising of the concentrations of dopamine,the temperature rise was more obvious,to a maximum of 30oC,the control group temperature was almost no fluctuate.The cancer cell capture capability with average efficiency of 84%.Adhesion effect and the release effect was very obviously,density slice layer and pattern layer test also quickly and ideal,showing its potential applications of the hydrogels.更多还原