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Digital Microfluidic Lab-on-a-Chip Platform for the Culture and Analysis of Three-Dimensional Multicellular Spheroids.

機譯：數(shù)字微流控芯片實驗室平臺，用于培養(yǎng)和分析三維多細胞球體。

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摘要

Cell spheroids are compact, multicellular aggregates grown in vitro that mimic the threedimensional morphology of in vivo tissues, thereby providing a more physiologically relevant tissue and disease model than cells cultured in two-dimensional monolayers. Despite the advantages of cell spheroids, their use in routine biomedical research has been limited, in part by the lack of automated, user-friendly, and flexible techniques for cell spheroid culture and analysis. Digital (droplet) microfluidics (DmuF) enables the manipulation of discrete drops of liquid through the controlled application of electric fields. Because it allows automated and flexible liquid handling, DmuF has the potential to address many of the shortcomings associated with current cell spheroid culture techniques. This dissertation describes the design, fabrication, and operation of a DmuF platform that enables the culture and analysis of three-dimensional, multicellular spheroids.;To enable spheroid culture on a DmuF device, a novel device architecture was developed that incorporates through-holes, or 'wells', in the bottom plate of the device that allow for the formation of hanging drops of controlled volume and composition. With the ability to create and address hanging drops in situ, protocols for automated cell spheroid culture were developed. Using DmuF liquid handling, spheroids can be initiated and maintained on the device for at least 96 hours, exhibiting good viability (>90%) and size uniformity (~8% CV intra-experiment, ~16% CV inter-experiment). Automated spheroid-based drug screening and migration assays were also performed, demonstrating the ability to create and assay spheroids with higher-order tissue properties as well as stimulate relevant physiological processes. An optimized DmuF peptide mass fingerprinting (PMF) sample preparation technique that could be useful for the in situ preparation and analysis of spheroid protein secretions was also developed.;The platform described here advances the field of digital microfluidics by introducing a novel functionality, hanging drop formation, which enables the culture of large, three-dimensional micro-tissues on a DmuF device. Improvements to the handling of biological solutions on a digital microfluidic device are also presented. This DmuF platform also represents an advance in the field of tissue engineering by providing a novel means of automating the culture and analysis of individually addressable cell spheroids. A DmuF platform that facilitates the culture and analysis of cell spheroids can potentially lower the barriers to adoption for the use of cell spheroids in routine biomedical research. Ultimately, broader use of spheroids in cell-based assays and screens has the potential to improve pre-clinical drug development efficiency and provide more physiologically relevant insights into our basic understanding of tissues and diseases.
機譯：細胞球體是體外生長的致密多細胞聚集體，可模仿體內(nèi)組織的三維形態(tài)，從而提供比在二維單層中培養(yǎng)的細胞更具生理相關(guān)性的組織和疾病模型。盡管細胞球體具有許多優(yōu)點，但它們在常規(guī)生物醫(yī)學研究中的使用受到了限制，部分原因是缺乏用于細胞球體培養(yǎng)和分析的自動化，用戶友好和靈活的技術(shù)。數(shù)字（液滴）微流控技術(shù)（DmuF）可以通過電場的受控施加來操縱離散的液滴。由于DmuF可以實現(xiàn)自動化和靈活的液體處理，因此有潛力解決與當前細胞球體培養(yǎng)技術(shù)相關(guān)的許多缺點。本文描述了DmuF平臺的設(shè)計，制造和操作，該平臺可進行三維多細胞球體的培養(yǎng)和分析。為了在DmuF器件上進行球體培養(yǎng)，開發(fā)了一種新型的器件結(jié)構(gòu)，該結(jié)構(gòu)結(jié)合了通孔，或“孔”，位于設(shè)備的底板中，可以形成受控制的體積和成分的懸掛液滴。具有在原位創(chuàng)建和處理懸滴的能力，開發(fā)了用于自動細胞球體培養(yǎng)的方案。使用DmuF液體處理，可以將球狀體初始化并在設(shè)備上維持至少96小時，表現(xiàn)出良好的生存能力（> 90％）和尺寸均勻性（?8％CV實驗內(nèi)，?16％CV實驗間）。還進行了基于球的自動藥物篩選和遷移分析，證明了創(chuàng)建和分析具有更高組織特性的球的能力以及刺激相關(guān)生理過程的能力。還開發(fā)了一種優(yōu)化的DmuF肽質(zhì)譜指紋圖（PMF）樣品制備技術(shù)，該技術(shù)可用于原位制備和球狀蛋白分泌的分析。;此處介紹的平臺通過引入一種新穎的功能，懸垂液滴來推進數(shù)字微流控領(lǐng)域。形成，這使得能夠在DmuF設(shè)備上培養(yǎng)大型三維微組織。還提出了對數(shù)字微流控設(shè)備上生物溶液處理的改進。該DmuF平臺還提供了一種自動化培養(yǎng)和分析單個可尋址細胞球體的新穎方法，從而代表了組織工程領(lǐng)域的一項進步。 DmuF平臺可促進細胞球的培養(yǎng)和分析，從而可以降低在常規(guī)生物醫(yī)學研究中采用細胞球的障礙。最終，在基于細胞的測定和篩選中更廣泛地使用球體，有可能提高臨床前藥物開發(fā)效率，并為我們對組織和疾病的基本理解提供更多與生理相關(guān)的見解。

著錄項

  • 作者

    Aijian, Andrew Peter.;

  • 作者單位

    University of California, Los Angeles.;

  • 授予單位 University of California, Los Angeles.;
  • 學科 Engineering Biomedical.
  • 學位 Ph.D.
  • 年度 2014
  • 頁碼 187 p.
  • 總頁數(shù) 187
  • 原文格式 PDF
  • 正文語種 eng
  • 中圖分類
  • 關(guān)鍵詞

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