Institute of Industrial Science, The University of Tokyo, Komaba 4-6-1, Meguro-ku, Tokyo 153-8505, Japan.
Lab Chip. 2014 Feb 21;14(4):730-6. doi: 10.1039/c3lc51086g.
Interest in the gene expression levels of pluripotent stem cells has increased in order to precisely understand cellular differentiation. Here, we propose a method utilizing a large number of arrayed microchambers to quantitatively measure an intracellular fluorescence protein that is genetically inserted to monitor a pluripotency marker protein, Nanog, in pluripotent stem cells. Individual cells are isolated and lysed by inducing an electric potential on the cell membrane within the tightly enclosed microchambers. The microchambers have a size that is comparable to the target cells, making it possible to trap single cells and restrict the dilution of the cell lysate. The amount of intracellular fluorescence proteins in a single cell is precisely quantified inside the well-defined volume of each microchamber. Our method will be a useful tool for high-throughput and parallelized read-outs of gene expression levels in individual cells in a large population of cells.
为了更精确地了解细胞分化，人们对多能干细胞的基因表达水平产生了浓厚的兴趣。在这里，我们提出了一种利用大量微室阵列来定量测量遗传插入的细胞内荧光蛋白的方法，以监测多能干细胞中的多能性标记蛋白 Nanog。通过在紧密封闭的微室中的细胞膜上施加电势，可以分离和裂解单个细胞。微室的大小与靶细胞相当，从而可以捕获单个细胞并限制细胞裂解物的稀释。在每个微室的明确定义的体积内，可以精确地定量单个细胞内的细胞内荧光蛋白的量。我们的方法将成为高通量和并行读取大量细胞中单个细胞基因表达水平的有用工具。
