Name: reconfigurable microfluidic channel with pin-discretized sidewalls
Uploaded: 2018-04-12T15:00:00.000+00:00
Duration: 10 min 39 s
Description: Watch this Scientific Journal Video about Reconfigurable Microfluidic Channel with Pin-discretized Sidewalls at JoVE.com

这项研究得到了 KAKENHI (20800048, 23700543) 的支持。

1. 针脚蚀刻 (图 2A)
<ol><li>脱脂在丙酮中浸泡的矩形销。</li>
	<li>通过浸泡在4毫升10% 硝酸, 然后加热溶液在65摄氏度为30分钟在烤箱钝化针。</li>
	<li>油脂实验在去离子水中引脚5分钟, 去除残留酸, 用纸巾擦干。将引脚浸入0.5 毫升的脱模剂中, 用于2小时油脂实验去离子水中的针脚5分钟。</li>
	<li>制造蚀刻盘 (图 2C)。<ol>
<li>使用标尺在玻璃滑轨上绘制或刻划两条平行线, 4 mm 间隙。</li>
		<li>对两个矩形切口盖玻片的一表面采用耐化学、低粘胶。</li>
		<li>债券的两个切割盖玻片在玻璃的幻灯片上, 在4毫米的差距。使用并行线作为参考。</li>
	</ol>
</li>
	<li>在蚀刻盘上分配两行硅胶粘合剂 (请参阅图 2C的位置和轮廓的大小)。 注: 3 维打印模板 (STL 模型文件作为补充材料包含) 将有助于轻松、准确地绘制线条。</li>
	<li>将针脚放在蚀刻盘上, 使其直端上的2毫米长的尖端浸入粘接线图案中。再次分配粘合剂, 以确保针脚完全覆盖并绘制轮廓。将蚀刻盘转移到加热到38摄氏度的湿润发酵罐中。在一夜之间治疗粘合剂。</li>
	<li>在玻璃瓶中缓慢添加0.2 毫升0.5 米硝酸到0.2 毫升的5.0 米盐酸。 注意: 混合物, 也称为水, 是非常腐蚀性和潜在的爆炸性。佩戴耐酸橡胶手套和安全护目镜, 在处理酸时要格外小心。永远不要存储解决方案。尽可能减少硝酸, 以减少其侵袭性。</li>
	<li>将蚀刻盘放在加热到65摄氏度的热板上。将酸混合物的0.4 毫升倒入针脚的未发现区域。等待10分钟, 将酸转移到烧杯中。</li>
	<li>用5毫升0.8 米碳酸氢钠溶液在去离子水中中和所有剩余的酸, 包括针脚的蚀刻区域。</li>
	<li>用镊子纵向拔出针脚, 从蚀刻盘中取出针脚。油脂实验在去离子水中的针脚为5分钟, 其次是丙酮超声波5分钟。</li>
	<li>钝化针脚的蚀刻区域, 如步骤1.2。</li>
	<li>用带十字线的车间显微镜检查蚀刻销的宽度。调整1.7 中描述的蚀刻时间, 使蚀刻区域的宽度为 0.2, 0.02 毫米。</li>
	<li>将针脚转移到含有5毫升70% 乙醇的玻璃瓶上。把瓶子带到层流罩上。把瓶子里的针脚拿出来, 让它们干。</li>
</ol>2. 制造含储层的硅酮板和一个销空间。
<ol><li>用典型的平版印刷工艺为销和固定侧壁制造一个空间模具。<ol>
<li>用1毫升的负环氧光刻胶涂上一脱脂玻璃滑梯, 使用旋转涂布机在1000转每分钟。将95摄氏度热板上的光刻胶干燥15分钟. 重复此步骤一次。</li>
		<li>自旋涂层的第三层相同的光刻胶在 2000 rpm 的玻璃幻灯片与涂层光刻胶。在95摄氏度热板上烘干30分钟的光刻胶。</li>
		<li>通过 photoplotted 薄膜将光刻胶层从 UV 光斑光源中暴露到450兆焦耳/cm2的 365 nm 紫外线。将暴露的光刻胶在95摄氏度热板上烘烤15分钟. 用手喷雾器喷涂溶剂 (2-甲氧基 1-丙基乙酸酯), 用氮气吹干, 研制出光刻胶。</li>
		<li>将玻璃滑梯与图案的光刻胶放在塑料盘子的底部。</li>
	</ol>
</li>
	<li>将烷的预聚体倒入模具上, 厚度为3毫米。Debubble 在真空干燥中的-800 帕预聚体为10分钟。</li>
	<li>用手术刀在65摄氏度的烤箱中治疗脱模1小时的预聚硅烷。在1小时内, 在120°c 的烤箱中完全治愈。</li>
	<li>沿着准则模式, 修剪不规则的边缘, 从与相同的手术刀板。尽可能精确和清洁剪切, 特别是在定义插入插槽的曲面 (请参见图 1a) 中的针脚。</li>
	<li>穿孔2毫米直径孔的入口/出口到主通道的两端, 使用活检拳。同样, 穿孔1毫米直径孔在排气通道的两端。有关通道布局和这些孔的位置, 请参见图 1A 。</li>
</ol>3. 装置的装配, 配有间隙填料和屏障的就地制造。
<ol><li>制造微通道组件。<ol>
<li>浸入 10 x 20 毫米 4 coverglass 在一个清洗解决方案加热到60°c 为10分钟。</li>
		<li>用去离子水冲洗盖玻片两次, 干燥120摄氏度, 10 分钟。</li>
		<li>等离子债券的 coverglass:<ol>
<li>在溅射涂布机的真空室中, 将该板 (通道功能侧上) 和清洗后的 10 x 20 mm 4 coverglass。</li>
			<li>开始抽水室到60宾夕法尼亚州产生空气真空等离子 (20 mA) 三十年代。</li>
			<li>立即发泄房间。将 coverglass 板的通道特征侧与边缘对齐。</li>
			<li>将粘合层放置在65摄氏度烤箱中, 10 分钟。</li>
		</ol>
</li>
		<li>使用无菌容器将粘合层带到层流罩上。用 UV 光消毒30分钟。</li>
		<li>在层流罩中, 将针脚插入插槽中, 使其两端形成微通道的其他侧壁。相邻的针脚长度应不同, 以避免两个垂直端的接触 (请参见图 1B)。垂直端之间的大间距是可取的。如果准备了不同 pin 长度 (L在图 2) 中的n类型的针脚, 则可能有 (N-1) x (pin 宽度) 的空间.</li>
	</ol>
</li>
	<li>制造一个基 (图 2B)。<ol>
<li>制作或读取底座的部件文件, 并使用 CAD/CAM 软件制作两个数控 (NC) 文件 (包含刀具路径; 包括辅助材料)。第一个补充 NC 文件使用4毫米直径的端铣刀和第二个1毫米直径的端铣刀。</li>
		<li>将3毫米厚的透明有机玻璃 (PMMA) 板夹在 CNC 轧机上。</li>
		<li>在计算机 NC (CNC) 轧机的控制器上打开第一个 nc 文件。将 4 mm 端铣刀安装到 CNC 铣削机上, 通过触摸端铣刀到 PMMA 板来定位零件零点。运行 NC 代码以剪切主板。 注: 偶尔用压缩空气吹干端铣刀以冷却和去除屑。</li>
		<li>使用第二个 NC 文件和 1 mm 端铣刀重复3.2.3。</li>
		<li>用洗涤剂脱脂加工部件, 用纸巾擦干。用70% 乙醇喷雾零件, 把它们带到层流罩里。</li>
	</ol>
</li>
	<li>制造针隙填料和弹性屏障: 注意: 步骤 3.3.1 3.3.7 应灌装在层流罩中执行。<ol>
<li>将白凡士林和聚四氟乙烯粉混合均匀, 按重量2:1 的比例制备间隙填料。使用超声波均质机融汇混合物。</li>
		<li>将缺口填料倒入注射器中。插入一个柱塞并将其推入注射器的尖端。连接针, 并再次推柱塞, 直到针尖被填补。同样, 用一个柱塞和一根针准备一个分配器注射器, 并填充硅胶粘合剂。</li>
		<li>使用适配器管将每个注射器连接到气动分配器。调整硅胶粘合剂和填料的分配压力为250帕和280帕。</li>
		<li>将硅胶粘合剂放在底部口袋的边缘。将 10 x 20 毫米 4 coverglass 放在口袋上, 并将其牢牢地压在键上。</li>
		<li>将硅胶粘合剂的深度约为1毫米, 沿底座的两个外槽绘制两段。将间隙填料分配到大约1毫米的深度, 以便沿另一个插槽绘制线段。</li>
		<li>将硅胶粘合剂放在另一口袋的边缘。将微通道组件 (3.1) 放在口袋上, 并将其牢固地压在键上。</li>
		<li>重复 3.3.5, 确保间隙填料和硅胶胶完全嵌入针脚, 并且插槽中没有开口。</li>
		<li>将该设备放入无菌容器中, 如带有盖子的不锈钢盒。将容器转移到加热到38摄氏度的湿润发酵罐中。在层流罩, 治愈的弹性屏障一天。</li>
		<li>将每个针脚沿相邻针脚移动1毫米, 以释放固化的弹性体屏障上的针脚。</li>
		<li>用 UV 光消毒设备30分钟。</li>
	</ol>
</li>
</ol>4. 微流控装置的评价
<ol><li>用荧光检测泄漏<ol>
<li>使用精细工具或台式机器人打开微通道。使通道宽度尽可能在整个通道中保持一致。</li>
		<li>将绿色荧光染料与去离子水稀释在10µM, 使荧光溶液。</li>
		<li>用 micropipettor 将荧光溶液添加到微通道的一端端口。此步骤将用解决方案填充通道。</li>
		<li>将微流控装置和两片吸水纸放在一个大的塑料盘子里, 湿用去离子水处理。在37摄氏度和 5% CO2上孵化出至少24小时的菜肴。</li>
		<li>用显微镜照相机用倒置荧光显微镜记录微通道的绿色荧光图像。</li>
		<li>使用适当的图像分析软件打开荧光图像, 并确认缝隙填料和针脚的接口上没有泄漏 (绿色荧光)。</li>
	</ol>
</li>
	<li>种子细胞到微通道。<ol>
<li>准备一个包含 70-80% 汇合细胞的细胞培养容器 (取决于细胞类型)。在生长培养基中分离并悬浮细胞。</li>
		<li>离心细胞 (速度和时间取决于细胞类型), 并吸入培养基。</li>
		<li>并用重悬有少量培养基的细胞。将单元格计数为单元格计数器, 并将单元格密度从 1.5 x 106调整为 1.5 x 107单元格/mL。</li>
		<li>使用精细工具或桌面机器人 (图 1B) 打开微通道, 以制作一个直400µm 的通道。调整针脚位置, 使侧壁尽可能平坦的整个通道。将单元格悬挂添加到微通道的一端端口之一, 并填充通道。</li>
		<li>找到定义区域的侧壁以开始区域性的针脚之一。在倒置显微镜下, 关闭两个相邻的针脚, 将细胞包围在细胞培养区内。</li>
		<li>从内到外关闭所有针脚以从通道中排出所有单元格。从端口中轻轻吸入悬浮液, 并向其添加介质。</li>
		<li>按照4.1.4 中描述的那样孵化设备。当细胞大约 70-80% 汇合, 慢慢地打开别针扩大文化区域。</li>
	</ol>
</li>
</ol>

<ol>
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作者声明他们没有竞争的财政利益。

引脚离散微通道是一种全功能的微流控通道, 我们认为与任何现有的微流控通道相比, reconfigurability 在通道形状上具有明显的高。我们在这里提供的协议将使细胞培养能够逐渐扩大细胞培养表面积的微流控装置能够长期保持融合下的培养。该装置还将在设计或制造时事先或任何其他考虑, 提供在基质中无图案蛋白质的细胞在通道内的图案。此外, 这种可重构微流控装置容易产生强大的通道位移流, 这将有助于实现处理这些难以流动的材料, 很少有现有的微流控设备可以处理。这意味着, 细胞和其他微生物、气体和其他非流体之间的相互作用可以使用该设备进行评估, 而不需要对设备设计进行大的修改。
我们考虑将拉普拉斯压力或静水压压力应用于通道的一个入口, 作为外部流量控制方法。我们不建议将液体推向死胡同, 因为它会通过针脚与通道的天花板/地板之间的缝隙产生流向排气通道的气流。许多流体操作不需要这样的 pin 操作。例如, 混合可以通过一个引脚 (即,只移动一个针来回几次) 的糖化液来完成。
设备最关键的部分是针脚。针脚需要精确的长度、平行度、垂直度和表面质量, 因为它们必须形成微通道, 必须平稳移动, 并且必须引导相邻针脚的移动。因此, 我们建议将针脚从专门进行精密加工的公司订购, 并提交类似于图 2a的绘图。可能有公司需要额外的几何尺寸和明确的表面粗糙度方向。但是, 如果这些别针是小心处理的, 偶尔用硝酸钝化, 这些针脚是可重用的。
弹性屏障是另一个关键特征, 其形成是器件制造过程中最关键的一步。为了获得可重复和可靠的结果, 需要一个精确加工的底座。在成灾屏障上放置针脚也是一个关键步骤。引脚应保持良好的对准, 并嵌入缝隙填料和屏障, 没有气泡。这些步骤防止泄漏通过引脚, 这是一个常见的问题, 这个微流控装置。
使用此设备的其他常见问题是: 摩擦约束引脚, b) 细胞死亡和低生长速率。可能的原因为这些在 a) 包括不均匀 (锥形或波浪) 蚀刻的针中间, 腐蚀表面的质量较差, 以及针尖端高度和光刻胶层高度之间的尺寸不匹配的硅板模具。调整蚀刻配方, 温度和搅拌可能有助于改善针运动。此外, 试件不使用蜡或胶粘剂将提供提示, 以解决问题。可能的因素在 b) 是没有足够的钝化的针脚, 在选择胶粘剂的弹性障碍, 和不完全固化胶粘剂。有些细胞可能需要在微通道内的涂层与纤维连接蛋白或其他蛋白质或聚合物, 促进细胞黏附。此外, 优化在细胞培养实践如 trypsinization 和离心将减少死细胞在微通道。
所提出的制造协议的一个局限性是, 只有一个壁被离散化。如果两个壁都是由 pin 阵列构建的, 则通道的 reconfigurability 将进一步提高。虽然它需要加倍的针脚数量和更长的制造步骤, 这是一个技术上可行的选择。

微流控装置-控制少量液体及其流动的微型设备--将生物医学程序小型化为 "芯片" 格式, 增加了可移植性, 而且往往是负担得起的。如最近的一次审查1所述, 开发了各种由空间和正则特性组成的微流控元件, 以实现混合、分离、微粒捕集或反应等基本和关键射流功能。
虽然许多微流控器件的行为是在设计阶段确定的, 但一些微流控装置允许对其结构或行为进行后期加工改变。在这里, 我们将此功能称为 "reconfigurability"。微流控系统的 reconfigurability 通常会减少设计设备所需的时间和成本, 并且/或能够随时定制微流控布局或功能。
以前描述的可重构微流控装置分为以下三类。首先, 弹性通道的变形允许在使用过程中改变流速和方向。为了获得 reconfigurability, 弹性通道由各种外部和可控制的力量变形, 如气压源2、盲文执行器3或压缩密封4。在第二种情况下, 可重构设备依赖于模块化设计, 如多层射流电路、带磁性互连的模块化通道和基于油管的微流体 5.第三, 设备本身不可重构, 但 microdroplet 在电极阵列上的传输 (通常称为数字微流体) 6、7和挂起式微流控装置8启用按需 流体的流动或路线的开关。
然而, 许多这些重新配置在拓扑和宏观水平上是有限的。例如, 许多集成的微流控设备通过在预定义区域中折叠微来停止流或改变流方向。但是, 要折叠的区域的位置和数量是不可重构的。虽然数字微流体具有多种流体处理能力, 但可能的流量应主要受每个液滴体积的限制。此外, 当细胞培养成这样的细胞培养基的水滴, 需要额外的努力, 以防止蒸发和气体消散的水滴, 避免渗透冲击和突然 pH 变化。
为了实现信道特征级 reconfigurability, 我们提出了一种具有移动壁的微流控装置, 它由机器元件阵列组成, 在使用9时可以动态地重新配置它们。为了形成一个可变形的侧壁, 小的长方形别针被排列了, 因此每个别针的末端定义了侧壁的部分。滑动的针脚允许的变形的侧壁, 允许运输或模式的细胞, 气泡, 和粒子在通道内。为了最小化死容量和最大化 reconfigurability, 相邻针脚之间的距离必须最小化。然而, 对连接在微通道内部和外部的针脚之间小间隙的强毛细管作用会导致任何液体进入针隙, 导致介质蒸发、细菌或细胞毒性污染, 最终细胞死亡。因此, 我们开发了无泄漏的离散侧壁型可重构微流控通道, 可以承受循环针动作和长期细胞培养10。
在这篇文章中, 我们提供了一个协议, 建立一个离散侧壁的微流化细胞培养装置, 可以重新配置后, 细胞培养区的逐步增加。利用荧光成像技术对离散通道壁的密封性进行了测试。利用片上细胞培养方法对细胞培养相容性和细胞模式能力进行评价。
这种微流控系统适用于任何适当的渠道设计不能预先确定, 必须改变的需求。例如, 该系统可用于根据细胞生长或迁移调整通道宽度和流速, 流动或诱捕活动线虫或其他在通道中异常行为的小物件, 或接受各种原始样品或生物制品,在设计时尚未构思。

<table><tbody><tr><td>Oven</td><td>Yonezawa</td><td>MI-100</td><td></td></tr><tr><td>10% Nitric Acid</td><td>Wako Chemicals</td><td>149-06845</td><td></td></tr><tr><td>Stainless steel pins</td><td>Micro Giken</td><td>N/A</td><td>0.3 mm crosssection, Grade 316L stainless steel, wire-cut EDM</td></tr><tr><td>Mold release agent</td><td>Fluoro Technology</td><td>FG-5093SH</td><td></td></tr><tr><td>Polydimethylsiloxane (PDMS)</td><td>Shin-Etsu Chemicals</td><td>KE-106</td><td></td></tr><tr><td>Negative epoxy photoresist</td><td>Nippon Kayaku</td><td>SU-8 3050</td><td></td></tr><tr><td>Coverglasses (Rectangular)</td><td>Matsunami Glass</td><td>26 x 60mm No.4</td><td></td></tr><tr><td>Acetone</td><td>Kanto Chemicals</td><td>01060-79</td><td></td></tr><tr><td>Glass slides (Large)</td><td>Matsunami Glass</td><td>76 x 52mm No.1</td><td></td></tr><tr><td>Silicone adhesive</td><td>Shin-Etsu Chemicals</td><td>KE-41</td><td></td></tr><tr><td>White petrolatum</td><td>Nikko Rica</td><td>Sun White P-1</td><td></td></tr><tr><td>Polytetrafluoroethylene (PTFE) powder</td><td>Power House Accele</td><td>Microfluon II</td><td></td></tr><tr><td>Clear acrylic plate (3 mm-thick)</td><td>Various</td><td>N/A</td><td></td></tr><tr><td>Pneumatic dispenser</td><td>Musashi Engineering</td><td>ML-5000XII</td><td></td></tr><tr><td>Hydrochloric acid</td><td>Kanto Chemicals</td><td>180768-00</td><td></td></tr><tr><td>Computer numerical control (CNC) mill</td><td>Pro Spec Tools</td><td>PSF240-CNC</td><td></td></tr><tr><td>End mill (4 mm diameter)</td><td>Mitsubishi Materials</td><td>MS2MSD0400</td><td></td></tr><tr><td>End mill (1 mm diameter)</td><td>Mitsubishi Materials</td><td>MS2MSD0100</td><td></td></tr><tr><td>Adhesive (chemical-resistant and low viscosity )</td><td>Cotronics</td><td>Duralco 4460</td><td></td></tr><tr><td>Borisilicate glass vials</td><td>Various</td><td></td><td>To prepare HNO3+HCl solution (Aqua regia). Always select borosilicate glass.</td></tr><tr><td>Sodium bicarbonate</td><td>Kanto Chemicals</td><td>37116-00</td><td></td></tr><tr><td>Ultrasonic cleaner</td><td>AS ONE</td><td>AS12GTU</td><td></td></tr><tr><td>Ultrasonic drill</td><td>Shinoda Tools</td><td>SOM-121</td><td>Used as a ultrasonic homogenizer.</td></tr><tr><td>Spin coater</td><td>Active</td><td>ACT-220DII</td><td></td></tr><tr><td>Hotplate</td><td>AS ONE</td><td>ND-1</td><td></td></tr><tr><td>Photoplotted film (12,700 dpi)</td><td>Unno Giken</td><td>N/A</td><td>Negative image of the recess at the bottom of a PDMS slab are plotted.</td></tr><tr><td>2-methoxy-1-methylethyl acetate</td><td>Wako Chemicals</td><td>130-10505</td><td></td></tr><tr><td>UV spot light source</td><td>Hamamatsu</td><td>L8327</td><td>Ultraviolet source</td></tr><tr><td>Nitrogen</td><td>Various</td><td>N/A</td><td></td></tr><tr><td>Vacuum desiccator and pump</td><td>AS ONE</td><td>MVD-100, GM-20S</td><td></td></tr><tr><td>Scalpels</td><td>Various</td><td>No.11</td><td></td></tr><tr><td>Biopsy punches (1.0mm and 2.0mm)</td><td>Kai Medical</td><td>BP-10F(1.0m), BP-20F(2.0mm)</td><td></td></tr><tr><td>Glass engraving pen</td><td>Various</td><td>N/A</td><td></td></tr><tr><td>Cleaning solution</td><td>Tama Chemicals</td><td>TMSC</td><td>Dilute 1:100 with deionized water</td></tr><tr><td>Sputter coater</td><td>San-yu Electron</td><td>SC-708</td><td>For plasma bonding.</td></tr><tr><td>Dispenser syringe (5 ml)</td><td>Musashi Engineering</td><td>PSY-5E</td><td></td></tr><tr><td>Plunger</td><td>Musashi Engineering</td><td>FLP-5E</td><td></td></tr><tr><td>Blunt needle (21G)</td><td>Musashi Engineering</td><td>PN-21G-B</td><td></td></tr><tr><td>Adapter tube</td><td>Musashi Engineering</td><td>AT-5E</td><td></td></tr><tr><td>Fermenter</td><td>Japan Kneader</td><td>PF100</td><td></td></tr><tr><td>Green fluorescent dye (Alexa Fluor 488 carboxylic acid)</td><td>Thermo Fisher</td><td>A33077</td><td></td></tr><tr><td>Large plastic dish</td><td>Greiner bio-one</td><td>688161</td><td></td></tr><tr><td>Absorbent paper</td><td>Asahi Kasei</td><td>BEMCOT M-1</td><td></td></tr><tr><td>Inverted microscope</td><td>Leica</td><td>DMi8</td><td></td></tr><tr><td>Microscope camera</td><td>Qimaging</td><td>Retiga 2000R</td><td></td></tr><tr><td>Dulbecco modified Eagle medium (DMEM)</td><td>GE Health Care</td><td>SH30021.01</td><td></td></tr><tr><td>Antibiotic-antimycotic solution</td><td>Thermo Fisher</td><td>15240-062</td><td></td></tr><tr><td>Trypsin/EDTA solution</td><td>Thermo Fisher</td><td>25200-056</td><td></td></tr><tr><td>Phosphate buffered saline (PBS)</td><td>GE Health Care</td><td>SH30256.01</td><td></td></tr><tr><td>Fetal bovine serum (FBS)</td><td>Biowest</td><td>S1820</td><td></td></tr><tr><td>Cell counter</td><td>FPI</td><td>OC-C-S02</td><td></td></tr><tr><td>Cell culture vessel</td><td>VIOLAMO</td><td>VTC-D100</td><td></td></tr><tr><td>15 ml conical tube</td><td>Corning</td><td>352095</td><td></td></tr><tr><td>Shop microscope</td><td>PEAK</td><td>2034-20</td><td></td></tr><tr><td>Hand sprayer</td><td>FURUPLA</td><td>No.3530</td><td></td></tr><tr><td>Coverglasses (Rectangular)</td><td>Matsunami Glass</td><td>10 x 20mm No.4</td><td></td></tr><tr><td>CAD/CAM software</td><td>Autodesk</td><td>Inventor HSM</td><td></td></tr><tr><td>Nitrogen gas pressure regulator</td><td>AS ONE</td><td>GF1-2506-RN-V</td><td>Set to 0.1 MPa</td></tr></tbody></table>

reconfigurable microfluidic channel with pin-discretized sidewalls

微流控元件需要有不同的形状, 以实现不同的关键微流控功能, 如混合, 分离, 粒子捕集, 或反应。在保持通道形状的情况下, 即使在制造过程中变形的微流控通道也能使高时空 reconfigurability。在现有的 "可重构" 或 "集成" 微流控系统中难以实现的关键微流控函数中, 需要这种 reconfigurability。我们描述了一种制造微流控通道的方法, 可变形侧壁由矩形销两端的侧向排列的阵列组成。在它们的纵向方向上驱动针脚会改变针脚的端点位置, 因此, 离散通道壁的形状。针隙可能导致不必要的泄漏或粘附在相邻的针脚造成的半月板力。为弥合引脚间隙, 我们引进了含烃类氟聚合物悬浮式缝隙填料, 并附有弹性屏障。这种可重构微流控装置可以产生强时间的通道内位移流, 也可以在通道的任何区域停止流动。这项功能将促进, 根据需要, 处理细胞, 粘性液体, 气泡和非流体, 即使他们的存在或行为是未知的, 在制造时。

可重构微通道的构造如图 1所示。在玻璃基板上放置了多个矩形销, 并将其排列起来, 使针脚的长边处于接触中。一张带有冲孔孔和与针脚高度相同深度的凹槽, 覆盖销的两端, 形成通道入口/出口储层, 通道天花板, 另一侧侧壁与由针脚组成的通道壁相反。该地区周围的针脚, 墙壁 (其中的一张面孔的一页), 玻璃基板形成一个微流控通道。
如前所述, 所提出的微流控系统的 reconfigurability 是由许多小的引脚平行放置在非常小但非零间隙。以前报告中的问题是通过毛细管效应产生的缝隙中的强流。为了克服这个问题, 缺口首先填补了缺口填料。在该协议中, 将粘性烃和含氟聚合物粉的分散混合物用作间隙填料。然而, 缝隙填料本身也受毛细管效应的影响。因此, 如图 1所示, 由此产生的可重构微通道既具有碳氢化合物/含氟聚合物间隙填料, 又有在间隙填料的外围周围形成的弹性体屏障。为了保证两个针脚之间的弹性屏障的厚度和强度, 需要将针脚的中间变薄, 以容纳足够数量的间隙填料。
图 2A显示形成侧边段的针脚的绘图。由于其耐腐蚀、浸出性能低, 选用不锈钢级316L 作为材料。然而, 需要额外的钝化过程, 使针细胞培养兼容。pin 必须有一个精确的矩形尖端没有毛刺, 以成功地形成一个侧壁段。此外, pin 必须有一个 "手柄", 以便通过推手柄可以很容易地移动 pin。由于每个引脚有一个狭窄的中间, 在别针之间的弹性体厚度足够承受由别针运动引起的剪切。与包括该装置的其他部件不同, 除中间细化外, 还应从专门从事电火花加工 (EDM) 的公司订购针脚, 因为它是加工小的最精确和经济高效的方法之一。由硬金属制成的零件。通过蚀刻来进行中间细化降低了加工成本和加工过程中弯曲或折断的风险。
为了确认间隙填料、弹性屏障和最终可重构微通道功能的防水, 采用荧光法进行泄漏检测。图 3显示了在微通道填充含荧光示踪染料的水后3天内, 弹性屏障边缘附近区域的荧光图像。荧光图像表明, 从弹性体屏障的可见边缘, 液体填充通道达到了约200µm 的深度。然而, 液体没有达到缝隙填料。此外, 没有发现缝隙填料通过弹性屏障的渗漏。这一观察表明, 狭窄的针中间和弹性屏障之间的紧密配合, 阻碍了液体通过缝隙的迁移。
最后, 我们通过逐步扩大可重构微流控装置的侧壁来进行长期细胞培养, 如图 4A所示。在0维, 少量的细胞被限制在一个等于一个针宽的空间内, 其他细胞也被吸气。在 2 d, 细胞被连接到底部表面并开始增殖。两个针脚被收回, 使所有细胞清晰可见, 虽然融合仍然低。在 5 d, 细胞继续增殖和融合增加。在6和 9 d, 其他两个针脚被收回, 以保持细胞 underconfluent。区域性的渐进扩展效果显示在图 4B中。在扣针的当天, 细胞密度发生了突变。然而, 细胞计数的生长速率保持不变, 而在典型细胞培养中看到的是指数。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/57230/57230fig1.jpg"> 图 1: 可重构微流控装置, 单针离散侧壁.(A)部件和构造可重构微流控装置。该装置有一个直通道与一个侧壁形成的两端10不锈钢引脚插入到单片机/玻璃微通道的特点。缝隙填料和弹性屏障防止液体通过针隙泄漏。盖玻片, 缝隙填料, 和弹性体屏障固定在一个有机玻璃 (PMMA) 基地。(B)自动 pin 机械手。一个由金属板制成的末端效应器被固定在3轴台式机器人上。要移动一个 pin, 最终效应器将推送其垂直端。不同长度的针脚放置在针脚宽度的三倍的间隔内。时间间隔可确保最终的效应器在一次有足够的间隙的情况下配对一个 pin。<a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/57230/57230fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/57230/57230fig2.jpg"> 图 2: 协议中使用的机械加工部件的力学图。单位以毫米;R指示半径维度; 否则为正方形符号 (-) 表示正方形特征;t表示粗细。(A) 316L 不锈钢引脚作为侧壁的一部分。可以按描述的顺序和加工针脚。针中间的细化使狗骨状形状不会在这幅图画中反映出来, 因为这不是作为加工的一部分被订购的, 而是作为协议的一部分进行的。(B)一种有机玻璃 (PMMA) 底座, 它持有盖玻片、缝隙填料和弹性屏障, 以防止针脚移动。(C)用于蚀刻针脚中间的蚀刻盘。为了制造蚀刻碟, 四块玻璃是用硅胶粘合剂粘合的。在盘子上画上了硅胶粘合剂的轮廓图案, 然后在盘子上放上针脚, 如图所示。<a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/57230/57230fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/57230/57230fig3.jpg"> 图 3: 通过 pin 间隙对可重构微通道的泄漏进行荧光检测.绿色荧光染料填充的荧光图像可重构微通道覆盖在密封结构的相衬图上, 该图像由间隙填料 (不透明) 和弹性体屏障 (半透明) 组成。弹性体屏障的边缘可见为半月板状特征, 并由上虚线表示;弹性体屏障与间隙填料之间的界面显示为与黑色区域接触的半月板状特征, 由下虚线表示。<a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/57230/57230fig3large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/57230/57230fig4.jpg"> 图 4: 可重构微通道中可变细胞培养区的渐进和连续细胞生长。(a)在细胞培养区内 COS-7 细胞生长, 受移动壁限制。(B)可重构微通道中受限的 COS-7 细胞密度的生长曲线和时间演化, 如下所示。三垂直箭头表示细胞培养区的扩展分别为2、5和 6 d。除了细胞计数, 细胞密度显示为相同的文化区域, 单独适合每个指数增长曲线, 并用于估计的局部加倍时间 (td [h]) 显示的框架。<a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/57230/57230fig4large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>

Watch this Scientific Journal Video about Reconfigurable Microfluidic Channel with Pin-discretized Sidewalls at JoVE.com

Reconfigurable Microfluidic Channel with Pin-discretized Sidewalls

一种具有可变形壁的微流控通道, 在使用时提供流量控制、粒子处理、信道维度定制等重新配置。我们描述了一种制造微流控通道的方法, 壁由一组引脚组成, 使其形状可以改变。

基于 Pin 离散壁的可重构微流控通道

Microfluidic components need to have various shapes to realize different key microfluidic functions such as mixing, separation, particle trapping, or ...

reconfigurable-microfluidic-channel-with-pin-discretized-sidewalls

Universidad San Sebastian

Research

JoVE Journal

Bioengineering

7.5K 次观看.  Shibaura Institute of Technology. 一种具有可变形壁的微流控通道, 在使用时提供流量控制、粒子处理、信道维度定制等重新配置。我们描述了一种制造微流控通道的方法, 壁由一组引脚组成, 使其形状可以改变。

视频: Reconfigurable Microfluidic Channel with Pin-discretized Sidewalls

A Microfluidic-based Hydrodynamic Trap for Single Particles

The ability to confine and manipulate single particles in free solution is a key enabling technology for fundamental and applied science. Methods for particle trapping based on optical, magnetic, electrokinetic, and acoustic techniques have led to major advancements in physics and biology ranging from the molecular to cellular level. In this article, we introduce a new microfluidic-based technique for particle trapping and manipulation based solely on hydrodynamic fluid flow. Using this method, we demonstrate trapping of micro- and nano-scale particles in aqueous solutions for long time scales. The hydrodynamic trap consists of an integrated microfluidic device with a cross-slot channel geometry where two opposing laminar streams converge, thereby generating a planar extensional flow with a fluid stagnation point (zero-velocity point). In this device, particles are confined at the trap center by active control of the flow field to maintain particle position at the fluid stagnation point. In this manner, particles are effectively trapped in free solution using a feedback control algorithm implemented with a custom-built LabVIEW code. The control algorithm consists of image acquisition for a particle in the microfluidic device, followed by particle tracking, determination of particle centroid position, and active adjustment of fluid flow by regulating the pressure applied to an on-chip pneumatic valve using a pressure regulator. In this way, the on-chip dynamic metering valve functions to regulate the relative flow rates in the outlet channels, thereby enabling fine-scale control of stagnation point position and particle trapping. The microfluidic-based hydrodynamic trap exhibits several advantages as a method for particle trapping. Hydrodynamic trapping is possible for any arbitrary particle without specific requirements on the physical or chemical properties of the trapped object. In addition, hydrodynamic trapping enables confinement of a "single" target object in concentrated or crowded particle suspensions, which is difficult using alternative force field-based trapping methods. The hydrodynamic trap is user-friendly, straightforward to implement and may be added to existing microfluidic devices to facilitate trapping and long-time analysis of particles. Overall, the hydrodynamic trap is a new platform for confinement, micromanipulation, and observation of particles without surface immobilization and eliminates the need for potentially perturbative optical, magnetic, and electric fields in the free-solution trapping of small particles.

In this article, we present a microfluidic-based method for particle confinement based on hydrodynamic flow. We demonstrate stable particle trapping at a fluid stagnation point using a feedback control mechanism, thereby enabling confinement and micromanipulation of arbitrary particles in an integrated microdevice.

一个单粒子为基础的微流控液力陷阱

The ability to confine and manipulate single particles in free solution is a key enabling technology for fundamental and applied science.  Methods for ...

科研

生物工程

Microfluidic Chips Controlled with Elastomeric Microvalve Arrays

Miniaturized microfluidic systems provide simple and effective solutions for low-cost point-of-care diagnostics and high-throughput biomedical assays. Robust flow control and precise fluidic volumes are two critical requirements for these applications. We have developed microfluidic chips featuring elastomeric polydimethylsiloxane (PDMS) microvalve arrays that: 1) need no extra energy source to close the fluidic path, hence the loaded device is highly portable; and 2) allow for microfabricating deep (up to 1 mm) channels with vertical sidewalls and resulting in very precise features.

The PDMS microvalves-based devices consist of three layers: a fluidic layer containing fluidic paths and microchambers of various sizes, a control layer containing the microchannels necessary to actuate the fluidic path with microvalves, and a middle thin PDMS membrane that is bound to the control layer. Fluidic layer and control layers are made by replica molding of PDMS from SU-8 photoresist masters, and the thin PDMS membrane is made by spinning PDMS at specified heights. The control layer is bonded to the thin PDMS membrane after oxygen activation of both, and then assembled with the fluidic layer. The microvalves are closed at rest and can be opened by applying negative pressure (e.g., house vacuum). Microvalve closure and opening are automated via solenoid valves controlled by computer software.

Here, we demonstrate two microvalve-based microfluidic chips for two different applications. The first chip allows for storing and mixing precise sub-nanoliter volumes of aqueous solutions at various mixing ratios. The second chip allows for computer-controlled perfusion of microfluidic cell cultures.

The devices are easy to fabricate and simple to control. Due to the biocompatibility of PDMS, these microchips could have broad applications in miniaturized diagnostic assays as well as basic cell biology studies.

We demonstrate protocols for manufacturing and automating elastomeric polydimethylsiloxane (PDMS)-based microvalve arrays that need no extra energy to close and feature photolithographically defined precise volumes. A parallel subnanoliter-volume mixer and an integrated microfluidic perfusion system are presented.

微流控芯片控制，弹性微阀阵列

Miniaturized microfluidic systems provide simple and effective solutions for low-cost point-of-care diagnostics and high-throughput biomedical assays.  ...

生物学

Microfluidic Pneumatic Cages: A Novel Approach for In-chip Crystal Trapping, Manipulation and Controlled Chemical Treatment

The precise localization and controlled chemical treatment of structures on a surface are significant challenges for common laboratory technologies. Herein, we introduce a microfluidic-based technology, employing a double-layer microfluidic device, which can trap and localize in situ and ex situ synthesized structures on microfluidic channel surfaces. Crucially, we show how such a device can be used to conduct controlled chemical reactions onto on-chip trapped structures and we demonstrate how the synthetic pathway of a crystalline molecular material and its positioning inside a microfluidic channel can be precisely modified with this technology. This approach provides new opportunities for the controlled assembly of structures on surface and for their subsequent treatment.

Herein, we describe the fabrication and operation of a double-layer microfluidic system made of polydimethylsiloxane (PDMS). We demonstrate the potential of this device for trapping, directing the coordination pathway of a crystalline molecular material and controlling chemical reactions onto on-chip trapped structures.

微流控气动笼:在芯片水晶捕获，处理的新方法和控制化学处理

The precise localization and controlled chemical treatment of structures on a surface are significant challenges for common laboratory technologies. ...

化学

Procedure for the Development of Multi-depth Circular Cross-sectional Endothelialized Microchannels-on-a-chip

Efforts have been focused on developing in vitro assays for the study of microvessels because in vivo animal studies are more time-consuming, expensive, and observation and quantification are very challenging. However, conventional in vitro microvessel assays have limitations when representing in vivo microvessels with respect to three-dimensional (3D) geometry and providing continuous fluid flow. Using a combination of photolithographic reflowable photoresist technique, soft lithography, and microfluidics, we have developed a multi-depth circular cross-sectional endothelialized microchannels-on-a-chip, which mimics the 3D geometry of in vivo microvessels and runs under controlled continuous perfusion flow. A positive reflowable photoresist was used to fabricate a master mold with a semicircular cross-sectional microchannel network. By the alignment and bonding of the two polydimethylsiloxane (PDMS) microchannels replicated from the master mold, a cylindrical microchannel network was created. The diameters of the microchannels can be well controlled. In addition, primary human umbilical vein endothelial cells (HUVECs) seeded inside the chip showed that the cells lined the inner surface of the microchannels under controlled perfusion lasting for a time period between 4 days to 2 weeks.

A microchannels-on-a-chip platform was developed by the combination of photolithographic reflowable photoresist technique, soft lithography, and microfluidics. The endothelialized microchannels platform mimics the three-dimensional (3D) geometry of in vivo microvessels, runs under controlled continuous perfusion flow, allows for high-quality and real-time imaging&#160;and can be applied for microvascular research.

发展多深的圆形横截面微内皮化的一个芯片的程序

Efforts have been focused on developing in vitro assays for the study of microvessels because in vivo animal studies are more time-consuming, expensive, ...

Microfluidic Fabrication of Polymeric and Biohybrid Fibers with Predesigned Size and Shape

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and used to dictate the shape as well as the diameter of a core stream.&#160;Grooves in the top and bottom of a microfluidic channel were designed to direct the sheath fluid and shape the core fluid.&#160;By matching the viscosity and hydrophilicity of the sheath and core fluids, the interfacial effects are minimized and complex fluid shapes can be formed.&#160;Controlling the relative flow rates of the sheath and core fluids determines the cross-sectional area of the core fluid.&#160;Fibers have been produced with sizes ranging from 300 nm to ~1 mm, and fiber cross-sections can be round, flat, square, or complex as in the case with double anchor fibers. Polymerization of the core fluid downstream from the shaping region solidifies the fibers.&#160;Photoinitiated click chemistries are well suited for rapid polymerization of the core fluid by irradiation with ultraviolet light.&#160;Fibers with a wide variety of shapes have been produced from a list of polymers including liquid crystals, poly(methylmethacrylate), thiol-ene and thiol-yne resins, polyethylene glycol, and hydrogel derivatives. Minimal shear during the shaping process and mild polymerization conditions also makes the fabrication process well suited for encapsulation of cells and other biological components.

Two adjacent fluids passing through a grooved microfluidic channel can be directed to form a sheath around a prepolymer core; thereby&#160;determining both shape and cross-section. Photoinitiated polymerization, such as thiol click chemistry, is well suited for rapidly solidifying the core fluid into a microfiber with predetermined size and shape.

预设计尺寸和形状的聚合物纤维和生物贿赂纤维的微流体制造

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and ...

One-Step Approach to Fabricating Polydimethylsiloxane Microfluidic Channels of Different Geometric Sections by Sequential Wet Etching Processes

Polydimethylsiloxane (PDMS) materials are substantially exploited to fabricate microfluidic devices by using soft lithography replica molding techniques. Customized channel layout designs are necessary for specific functions and integrated performance of microfluidic devices in numerous biomedical and chemical applications (e.g., cell culture, biosensing, chemical synthesis, and liquid handling). Owing to the nature of molding approaches using silicon wafers with photoresist layers patterned by photolithography as master molds, the microfluidic channels commonly have regular cross sections of rectangular shapes with identical heights. Typically, channels with multiple heights or different geometric sections are designed to possess particular functions and to perform in various microfluidic applications (e.g., hydrophoresis is used for sorting particles and in continuous flows for separating blood cells6,7,8,9). Therefore, a great deal of effort has been made in constructing channels with various sections through multiple-step approaches like photolithography using several photoresist layers and assembly of different PDMS thin sheets. Nevertheless, such multiple-step approaches usually involve tedious procedures and extensive instrumentation. Furthermore, the fabricated devices may not perform consistently and the resulted experimental data may be unpredictable. Here, a one-step approach is developed for the straightforward fabrication of microfluidic channels with different geometric cross sections through PDMS sequential wet etching processes, that introduces etchant into channels of planned single-layer layouts embedded in PDMS materials. Compared to the existing methods for manufacturing PDMS microfluidic channels with different geometries, the developed one-step approach can significantly simplify the process to fabricate channels with non-rectangular sections or various heights. Consequently, the technique is a way of constructing complex microfluidic channels, which provides a fabrication solution for the advancement of innovative microfluidic systems.

Several methods are available for the fabrication of channels of non-rectangular sections embedded in polydimethylsiloxane microfluidic devices. Most of them involve multistep manufacturing and extensive alignment. In this paper, a one-step approach is reported for fabricating microfluidic channels of different geometric cross sections by polydimethylsiloxane sequential wet etching.

连续湿法刻蚀法制作不同几何断面烷微流控通道的一步法

Polydimethylsiloxane (PDMS) materials are substantially exploited to fabricate microfluidic devices by using soft lithography replica molding techniques. ...

工程学

Multi-step Variable Height Photolithography for Valved Multilayer Microfluidic Devices

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to entry for non-specialists who would benefit greatly from the ability to develop their own microfluidic devices to address research questions. Particularly lacking has been the open dissemination of protocols related to photolithography, a key step in the development of a replica mold for the manufacture of polydimethylsiloxane (PDMS) devices. While the fabrication of single height silicon masters has been explored extensively in literature, fabrication steps for more complicated photolithography features necessary for many interesting device functionalities (such as feature rounding to make valve structures, multi-height single-mold patterning, or high aspect ratio definition) are often not explicitly outlined. 
Here, we provide a complete protocol for making multilayer microfluidic devices with valves and complex multi-height geometries, tunable for any application. These fabrication procedures are presented in the context of a microfluidic hydrogel bead synthesizer and demonstrate the production of droplets containing polyethylene glycol (PEG diacrylate) and a photoinitiator that can be polymerized into solid beads. This protocol and accompanying discussion provide a foundation of design principles and fabrication methods that enables development of a wide variety of microfluidic devices. The details included here should allow non-specialists to design and fabricate novel devices, thereby bringing a host of recently developed technologies to their most exciting applications in biological laboratories.

Multilayer microfluidic devices often involve the fabrication of master molds with complex geometries for functionality. This article presents a complete protocol for multi-step photolithography with valves and variable height features tunable to any application. As a demonstration, we fabricate a microfluidic droplet generator capable of producing hydrogel beads.

多级可变高度的光刻多层带瓣微流体装置

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to ...

Soft Lithographic Procedure for Producing Plastic Microfluidic Devices with View-ports Transparent to Visible and Infrared Light

Infrared (IR) spectro-microscopy of living biological samples is hampered by the absorption of water in the mid-IR range and by the lack of suitable microfluidic devices. Here, a protocol for the fabrication of plastic microfluidic devices is demonstrated, where soft lithographic techniques are used to embed transparent Calcium Fluoride (CaF2) view-ports in connection with observation chamber(s). The method is based on a replica casting approach, where a polydimethylsiloxane (PDMS) mold is produced through standard lithographic procedures and then used as the template to produce a plastic device. The plastic device features ultraviolet/visible/infrared (UV/Vis/IR) -transparent windows made of CaF2 to allow for direct observation with visible and IR light. The advantages of the proposed method include: a reduced need for accessing a clean room micro-fabrication facility, multiple view-ports, an easy and versatile connection to an external pumping system through the plastic body, flexibility of the design, e.g., open/closed channels configuration, and the possibility to add sophisticated features such as nanoporous membranes.

A protocol for the fabrication of plastic microfluidic devices with transparent view-ports for visible and infrared light imaging is described.

生产塑料微流控装置与查看端口对可见光和红外透明软光刻过程

Infrared (IR) spectro-microscopy of living biological samples is hampered by the absorption of water in the mid-IR range and by the lack of suitable ...

Generation of Dynamical Environmental Conditions using a High-Throughput Microfluidic Device

Mimicking in vivo environmental conditions is crucial for in vitro studies on complex life machinery. However, current techniques targeting live cells and organs are either highly expensive, like robotics, or lack nanoliter volume and millisecond time accuracy in liquid manipulation. We herein present the design and fabrication of a microfluidic system, which consists of 1,500 culture units, an array of enhanced peristaltic pumps and an on-site mixing modulus. To demonstrate the capacities of the microfluidic device, neural stem cell (NSC) spheres are maintained in the proposed system. We observed that when the NSC sphere is exposed to CXCL in day 1 and EGF in day 2, the round-shaped conformation is well maintained. Variation in the input order of 6 drugs causes morphological changes to the NSC sphere and the expression level representative marker for NSC stemness (i.e., Hes5 and Dcx). These results indicate that dynamic and complex environmental conditions have great effects on NSC differentiation and self-renewal, and the proposed microfluidic device is a suitable platform for high throughput studies on the complex life machinery.

We present a microfluidic system for high throughput studies on complex life machinery, which consists of 1500 culture units, an array of enhanced peristaltic pumps and an on-site mixing modulus. The microfluidic chip allows for the analysis of the highly complex and dynamic micro-environmental conditions in vivo.

使用高通量微流体设备生成动态环境条件

Mimicking in vivo environmental conditions is crucial for in vitro studies on complex life machinery. However, current techniques targeting live cells and ...

用于组合插头生产的微流体装置的设置

Bilayer Microfluidic Device for Combinatorial Plug Production

Droplet microfluidics is a versatile tool that allows the execution of a large number of reactions in chemically distinct nanoliter compartments. Such systems have been used to encapsulate a variety of biochemical reactions - from incubation of single cells to implementation of PCR reactions, from genomics to chemical synthesis. Coupling the microfluidic channels with regulatory valves allows control over their opening and closing, thereby enabling the rapid production of large-scale combinatorial libraries consisting of a population of droplets with unique compositions. In this paper, protocols for the fabrication and operation of a pressure-driven, PDMS-based bilayer microfluidic device that can be utilized to generate combinatorial libraries of water-in-oil emulsions called plugs are presented. By incorporating software programs and microfluidic hardware, the flow of desired fluids in the device can be controlled and manipulated to generate combinatorial plug libraries and to control the composition and quantity of constituent plug populations. These protocols will expedite the process of generating combinatorial screens, particularly to study drug response in cells from cancer patient biopsies.

作者聚焦：在精准肿瘤学中整合计算和实验方法

The fabrication of a polydimethylsiloxane (PDMS)-based bilayer device for the production of combinatorial libraries in water-in-oil emulsions (plugs) is presented here. The necessary hardware and software required to automate plug production are detailed in the protocol, and the production of a quantitative library of fluorescent plugs is also demonstrated.

用于组合塞生产的双层微流控装置

Droplet microfluidics is a versatile tool that allows the execution of a large number of reactions in chemically distinct nanoliter compartments. Such ...