Expanding the Available Assays: Adapting and Validating In-Cell Westerns in Microfluidic Devices for Cell-Based Assays
Expanding the Available Assays: Adapting and Validating In-Cell Westerns in Microfluidic Devices for Cell-Based Assays
复制标题
DOI：
10.1089/adt.2010.0274
复制
发表时间：
2010-10-01
影响因子：
1.8
通讯作者：
Beebe, David J.
中科院分区：
文献类型：
--
作者：
Paguirigan, Amy L.;Puccinelli, John P.;Beebe, David J.
与许多传统的培养技术相比，用于细胞研究的微流体方法可以显著降低成本，因为它减少了试剂和生物标本的需求。然而，目前的读数类型是有限的，缺乏适合微流体培养的读数，这极大地阻碍了微流体在细胞基础分析中的应用。细胞Western （ICW）技术使用定量免疫细胞化学和激光扫描仪来提供在任意几何形状的微流体通道中生长的细胞中蛋白质数量的原位测量。通过与当前宏观尺度方法的详细比较，验证了ICWs在微流控通道中的应用，并显示出良好的相关性。以转化生长因子β诱导上皮细胞系向间充质转化为例，进一步验证了该技术在高通量微流控通道中作为基于可溶性因子的检测的读数。使用被动泵送样品和激光扫描进行分析打开了高通量定量微流体细胞为基础的分析，与现有的高通量基础设施无缝集成的大门。
Microfluidic methods for cellular studies can significantly reduce costs due to reduced reagent and biological specimen requirements compared with many traditional culture techniques. However, current types of readouts are limited and this lack of suitable readouts for microfluidic cultures has significantly hindered the application of microfluidics for cell-based assays. The In- Cell Western (ICW) technique uses quantitative immunocytochemistry and a laser scanner to provide an in situ measure of protein quantities in cells grown in microfluidic channels of arbitrary geometries. The use of ICWs in microfluidic channels was validated by a detailed comparison with current macroscale methods and shown to have excellent correlation. Transforming growth factor-beta-induced epithelial-to-mesenchymal transition of an epithelial cell line was used as an example for further validation of the technique as a readout for soluble-factor-based assays performed in high-throughput microfluidic channels. The use of passive pumping for sample delivery and laser scanning for analysis opens the door to high-throughput quantitative microfluidic cell-based assays that integrate seamlessly with existing high-throughput infrastructure.