Microfluidic device for DNA amplification of single cancer cells isolated from whole blood by self-seeding microwells
Microfluidic device for DNA amplification of single cancer cells isolated from whole blood by self-seeding microwells
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DOI：
10.1039/c5lc00816f
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发表时间：
2015-01-01
期刊：
影响因子：
6.1
通讯作者：
Terstappen, Leon W. M. M.
中科院分区：
文献类型：
--
作者：
Yang, Yoonsun;Rho, Hoon Suk;Terstappen, Leon W. M. M.
自接种微孔芯片可以基于细胞大小将单细胞分选到6400个威尔斯孔中，并通过免疫荧光染色验证其身份。在这里，我们开发了一种微流体装置，可以将这些单细胞放置在其中，裂解并扩增其DNA以进行进一步的询问。在白细胞去除后，使掺入MCF7肿瘤细胞的全血通过微孔芯片，并且通过微孔中的上皮细胞粘附分子（EpCAM）染色鉴定37%的MCF7细胞。将鉴定的单细胞冲压到微流体装置的反应室中，并通过微阀的蠕动泵送顺序引入用于细胞裂解和DNA扩增的试剂。在8个平行室中进行芯片上裂解和扩增，产生10000倍的DNA扩增。样品通过反应室的可接近性允许容易地检索和询问靶特异性基因以表征肿瘤细胞。
Self-seeding microwell chips can sort single cells into 6400 wells based on cell size and their identity verified by immunofluorescence staining. Here, we developed a microfluidic device in which these single cells can be placed, lysed and their DNA amplified for further interrogation. Whole blood spiked with MCF7 tumor cells was passed through the microwell chips after leukocyte depletion and 37% of the MCF7 cells were identified by epithelial cell adhesion molecule ( EpCAM) staining in the microwells. Identified single cells were punched into the reaction chamber of the microfluidic device and reagents for cell lysis and DNA amplification introduced sequentially by peristaltic pumping of micro-valves. On-chip lysis and amplification was performed in 8 parallel chambers yielding a 10000 fold amplification of DNA. Accessibility of the sample through the reaction chamber allowed for easy retrieval and interrogation of target-specific genes to characterize the tumor cells.