Continuous cell introduction for the analysis of individual cells by capillary electrophoresis
Continuous cell introduction for the analysis of individual cells by capillary electrophoresis
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DOI：
10.1021/ac0009088
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发表时间：
2001-01-01
影响因子：
7.4
通讯作者：
Lillard, SJ
中科院分区：
文献类型：
--
作者：
Chen, SJ;Lillard, SJ
描述了用于通过毛细管电泳对单细胞进行高通量分析的仪器。使用电渗流（electroosmotic now，电渗流）的基于流动的界面提供了完整细胞通过引入毛细管到细胞裂解接合部中的连续注射以及所得细胞裂解物通过分离毛细管的迁移以用于分析。具体地说，两个毛细管用5毫米长的特氟隆管连接在一起，以产生类似于5 μ m的间隙，并将连接处浸入缓冲液储存器中。在两个毛细管上施加高电压，使得细胞通过离心连续泵入第一个毛细管中。在没有洗涤剂的情况下，单个细胞在连接处在柱上裂解，推测是由于差距处的NOW性质的急剧变化引起的机械破坏。每个细胞在连接处裂解后，通过毛细管电泳分离主要蛋白质血红蛋白和碳酸酐酶，并使用275 nm激发通过激光诱导天然荧光检测所得分析物区域。血红蛋白和碳酸酐酶的检出限分别为37和1.6-阿莫尔，与文献值吻合良好。使用完整的红细胞对器械进行了评价。完成分析的平均时间（即，连续注射、裂解、分离和检测）小于4分钟。此外，该仪器简化了单个完整细胞的引入，而无需使用显微镜。
Instrumentation for high-throughput analysis-of single cells by capillary electrophoresis is described, A flow-based; interface that uses electroosmotic now (EOF) provides continuous injection of intact cells through an introduction capillary into a cell lysis junction and migration of the resulting cell lysate through a separation capillary for analysis. Specifically two capillaries-were coupled together with 5-mm-long Teflon tubing to create a similar to5-mum gap, and the junction was immersed in a buffer reservoir. High voltage was applied-across both capillaries so that cells were continuously pumped into the first capillary by EOF, Individual cells were lysed on-column at the junction without detergents, presumably owing to mechanical disruption caused by a dramatic change in now properties at the gap. After each cell was lysed at the junction, the major proteins hemoglobin and carbonic anhydrase were separated by capillary electrophoresis and the resultant analyte zones were detected by laser induced native fluorescence using 275-nm excitation. The detection limits of hemoglobin and carbonic anhydrase were 37 and 1.6-amol, respectively, which correlate well with the literature. The instrumentation was evaluated with intact red: blood cells. The averaged time for complete analysis (i.e., continuous injection, lysis, separation,:and detection) of one human erythrocyte was less than 4 min with this capillary-based setup, Moreover, this instrumentation simplifies the introduction of individual; intact cells without the use of a microscope.