發(fā)布時(shí)間：2018-04-07 19:04

  本文選題：液滴鏈　切入點(diǎn)：液滴微流控　出處：《浙江大學(xué)》2016年博士論文

【摘要】：構(gòu)建體外細(xì)胞遷移和共培養(yǎng)模型將有利于藥物篩選、早期疾病診斷和基礎(chǔ)生物學(xué)的研究。微流控系統(tǒng)在芯片結(jié)構(gòu)的設(shè)計(jì)、細(xì)胞培養(yǎng)的材料、流體控制的方法上都具備多種選擇,可以用于細(xì)胞微環(huán)境的控制和模擬,實(shí)現(xiàn)細(xì)胞三維培養(yǎng)和共培養(yǎng),進(jìn)而用于研究細(xì)胞行為、探究病理生理現(xiàn)象、開發(fā)和篩選新的藥物。然而大多用于細(xì)胞遷移行為和共培養(yǎng)研究的微流控裝置受芯片復(fù)雜性和流體控制裝置的限制,難以適用于多種細(xì)胞生物學(xué)研究。本工作的目的是建立一個(gè)簡單的多模式液滴微流控方法,利用基于多孔膜的液滴鏈裝置完成多種細(xì)胞實(shí)驗(yàn),用于低消耗、高通量的細(xì)胞生物學(xué)研究,包括多模式細(xì)胞遷移的研究和多細(xì)胞共培養(yǎng)模型的建立。第一章我們綜述了應(yīng)用于細(xì)胞遷移、細(xì)胞三維培養(yǎng)、細(xì)胞共培養(yǎng)的微流控芯片技術(shù)的研究進(jìn)展。物質(zhì)濃度梯度的形成對于細(xì)胞遷移的研究至關(guān)重要,我們主要分類介紹了基于多相層流、微通道、凝膠、多孔膜體系等構(gòu)建的濃度梯度生成系統(tǒng)及其在細(xì)胞遷移實(shí)驗(yàn)中的應(yīng)用。在本章中,我們還介紹了微流控三維細(xì)胞培養(yǎng)的主要方法,包括利用懸滴和凝膠等方法實(shí)現(xiàn)細(xì)胞的三維生長；細(xì)胞共培養(yǎng)的方法則主要介紹了圖案化的細(xì)胞共培養(yǎng)方法和基于膜的與基于通道的細(xì)胞共培養(yǎng)方法；以及利用細(xì)胞三維培養(yǎng)、共培養(yǎng)技術(shù)構(gòu)建生理相關(guān)的多器官組織模型的進(jìn)展。在第二章中,我們建立了一種基于多孔膜的三維液滴鏈陣列系統(tǒng)用于多模式細(xì)胞遷移的研究。三維液滴鏈通過多孔膜上的微孔進(jìn)行物質(zhì)交流,通過PDMS通孔陣列片控制膜上下液滴的相對位置,改變液滴的體積、位置、內(nèi)容物可以控制物質(zhì)擴(kuò)散速度,生成不同的濃度梯度,進(jìn)行多種模式的細(xì)胞遷移實(shí)驗(yàn)。液滴鏈中液滴的體積在500-800 nL之間,一個(gè)芯片上可完成高達(dá)81個(gè)細(xì)胞遷移實(shí)驗(yàn)。我們將這一細(xì)胞遷移系統(tǒng)應(yīng)用于多種模式的細(xì)胞遷移實(shí)驗(yàn)中,包括精準(zhǔn)的細(xì)胞遷移實(shí)驗(yàn)、競爭性細(xì)胞遷移實(shí)驗(yàn)、仿生趨化性細(xì)胞遷移實(shí)驗(yàn)。在第三章中,基于第二章的方法,我們設(shè)計(jì)了集成度更高的花型液滴鏈和珠串型液滴鏈芯片。在花型液滴鏈中,將懸滴三維培養(yǎng)、多細(xì)胞共培養(yǎng)的方法用于多種細(xì)胞相互作用的研究中,構(gòu)建了集成肝、肺、腎、腸、心等細(xì)胞的多器官液滴鏈陣列芯片,并應(yīng)用于轉(zhuǎn)移性乳腺癌細(xì)胞MDA-MB-231向不同器官微組織的遷移傾向性研究。初步構(gòu)建了基于生理?xiàng)l件建立的珠串型液滴鏈芯片,以模擬藥物在體內(nèi)的代謝過程。
[Abstract]:The establishment of cell migration and co-culture model in vitro will be beneficial to drug screening, early disease diagnosis and basic biology research.The microfluidic system has many choices in chip structure design, cell culture materials and fluid control methods. It can be used to control and simulate cell microenvironment and realize cell three-dimensional culture and co-culture.It can be used to study cell behavior, explore pathophysiological phenomena, and develop and screen new drugs.However, most of the microfluidic devices used in cell migration and co-culture are limited by chip complexity and fluid control devices, which are difficult to be applied to many cell biology studies.The purpose of this work is to establish a simple multi-mode droplet microfluidic method, and to use the liquid droplet chain device based on porous membrane to complete a variety of cell experiments, which can be used in low consumption and high throughput cell biology research.Including the study of multi-model cell migration and the establishment of multi-cell co-culture model.In the first chapter, we review the research progress of microfluidic chip technology used in cell migration, cell three-dimensional culture and cell co-culture.The formation of concentration gradient is very important to the study of cell migration. We mainly introduce the concentration gradient generation system based on multiphase laminar flow, microchannel, gel and porous membrane system and its application in cell migration experiments.In this chapter, we also introduce the main methods of microfluidic three-dimensional cell culture, including the use of suspension and gel methods to achieve three-dimensional cell growth;The methods of cell coculture mainly include patterned cell coculture, membrane-based and channel-based co-culture, and three-dimensional cell culture.Progress of co-culture techniques in the construction of physiological-related multi-organ tissue models.In the second chapter, we establish a three-dimensional droplet chain array system based on porous membrane for the study of multi-mode cell migration.The three-dimensional droplet chain communicates through the micropores in the porous membrane, and the relative position of the droplets up and down the membrane is controlled by the PDMS through pore array. The volume, position and contents of the droplets can control the diffusion velocity of the material and produce different concentration gradients.Cell migration experiments were carried out in a variety of models.The volume of droplets in the droplet chain ranges from 500 to 800 NL, and up to 81 cell migration experiments can be performed on a single chip.We applied this cell migration system to many cell migration experiments, including accurate cell migration, competitive cell migration, and bionic chemotactic cell migration.In Chapter 3, based on the method in Chapter 2, we design a more integrated droplet chain and a droplet chain chip.In the floral droplet chain, the method of suspension three-dimensional culture and multicellular co-culture was used in the study of multi-cell interaction, and a multi-organ droplet chain array of liver, lung, kidney, intestine, heart and so on was constructed.It was used to study the migration tendency of metastatic breast cancer cell line MDA-MB-231 to different organ microtissues.A series of droplet chain chips based on physiological conditions were constructed to simulate the drug metabolism in vivo.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：Q813

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本文編號：1720475