Fluorescence-activated droplet sorting (FADS): efficient microfluidic cell sorting based on enzymatic activity
Fluorescence-activated droplet sorting (FADS): efficient microfluidic cell sorting based on enzymatic activity
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DOI：
10.1039/b902504a
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发表时间：
2009-01-01
期刊：
影响因子：
6.1
通讯作者：
Griffiths, Andrew D.
中科院分区：
文献类型：
--
作者：
Baret, Jean-Christophe;Miller, Oliver J.;Griffiths, Andrew D.
我们描述了一个高效的微流体荧光激活的液滴分散器（FADS），结合了微构板筛选和传统荧光激活的细胞分选（FACS）的许多优势。单细胞在乳液液滴中被划分，可以以荧光激活的方式（如FACS）以直至2000滴S（-1）的速率对其进行分类。为了验证该系统，用荧光底物对表达报告酶β-半乳糖苷酶或非活性变体的大肠杆菌细胞的混合物进行了分际划分，并以类似于300滴S（-1）的速率分选。该吞吐量在此吞吐量下的误差误差率在10（4）个液滴中<1。对分类细胞的分析表明，富集的主要极限是大肠杆菌细胞的共囊化，而不是分类误差：基于泊松分布的理论模型准确地预测了使用起始细胞密度（每个液滴细胞）观察到的富集值（ ）以及活性细胞的比率。当细胞以低密度封装（每50滴类似于1个细胞）时，排序非常有效，所有回收的细胞都是活性应变。另外，对单个活性液滴进行了分类，并成功恢复了细胞。
We describe a highly efficient microfluidic fluorescence-activated droplet sorter (FADS) combining many of the advantages of microtitre-plate screening and traditional fluorescence-activated cell sorting (FACS). Single cells are compartmentalized in emulsion droplets, which can be sorted using dielectrophoresis in a fluorescence-activated manner (as in FACS) at rates up to 2000 droplets s(-1). To validate the system, mixtures of E. coli cells, expressing either the reporter enzyme beta-galactosidase or an inactive variant, were compartmentalized with a fluorogenic substrate and sorted at rates of similar to 300 droplets s(-1). The false positive error rate of the sorter at this throughput was < 1 in 10(4) droplets. Analysis of the sorted cells revealed that the primary limit to enrichment was the co-encapsulation of E. coli cells, not sorting errors: a theoretical model based on the Poisson distribution accurately predicted the observed enrichment values using the starting cell density (cells per droplet) and the ratio of active to inactive cells. When the cells were encapsulated at low density (similar to 1 cell for every 50 droplets), sorting was very efficient and all of the recovered cells were the active strain. In addition, single active droplets were sorted and cells were successfully recovered.