Institute of Industrial Science, The University of Tokyo, 4-6-1, Komaba, Meguro-ku, Tokyo 153-8505, Japan.
Small. 2011 Nov 18;7(22):3239-47. doi: 10.1002/smll.201101028. Epub 2011 Sep 20.
We present a novel method, implemented in the form of a microfluidic device, for arraying and analyzing large populations of single cells. The device contains a large array of electroactive microwells where manipulation and analysis of large population of cells are carried out. On the device, single cells can be actively trapped in the microwells by dielectrophoresis (DEP) and then lysed by electroporation (EP) for subsequent analysis of the confined cell lysates. The DEP force in the selected dimensions of the microwells could achieve efficient trapping in nearly all the microwells (95%) in less than three minutes. Moreover, the positions of the cells in the microwells are maintained even when unstable flow of liquid is applied. This makes it possible to exchange the DEP buffer to a solution that will be subsequently used for stimulating or analyzing the trapped cells. After closing the microwells, EP is conducted to lyse the trapped cells by applying short electric pulses. Tight enclosure is critical to prevent dilution, diffusion and cross contamination of the cell lysates. We demonstrated the feasibility of our approach with an enzymatic assay measuring the intracellular-galactosidase activity. The use of this method should greatly help analysis of large populations of cells at the single-cell level. Furthermore, the method offers rapidity in the trapping and analysis of multiple cell types in physiological conditions that will be important to ensure the relevance of single cell analyses.
我们提出了一种新的方法,以微流控装置的形式实现,用于对大量单细胞进行阵列和分析。该装置包含一个大型的电活性微井阵列,可在此对大量细胞进行操作和分析。在该装置上,通过介电泳(DEP)可以主动将单细胞捕获在微井中,然后通过电穿孔(EP)将其裂解,以便随后分析封闭的细胞裂解物。在微井的选定尺寸中,DEP 力可以在不到三分钟的时间内将几乎所有微井(95%)中的细胞有效捕获。此外,即使施加不稳定的液体流动,微井中细胞的位置也能保持不变。这使得可以将 DEP 缓冲液交换为随后用于刺激或分析捕获细胞的溶液。封闭微井后,通过施加短电脉冲进行 EP 以裂解捕获的细胞。紧密的封闭对于防止细胞裂解物的稀释、扩散和交叉污染至关重要。我们通过测量细胞内半乳糖苷酶活性的酶联免疫吸附测定法验证了我们方法的可行性。该方法的使用将极大地有助于单细胞水平上对大量细胞进行分析。此外,该方法在生理条件下具有快速捕获和分析多种细胞类型的能力,这对于确保单细胞分析的相关性非常重要。