Concurrent isolation of lymphocytes and granulocytes using prefocused free flow acoustophoresis.
Concurrent isolation of lymphocytes and granulocytes using prefocused free flow acoustophoresis.
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DOI：
10.1021/acs.analchem.5b00370
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发表时间：
2015-06-02
影响因子：
7.4
通讯作者：
Laurell T
中科院分区：
文献类型：
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作者：
Grenvall C;Magnusson C;Lilja H;Laurell T
基于微芯片的自由流声泳（FFA）结合二维细胞预聚焦，使并行多目标出口分馏的白细胞成亚群（淋巴细胞，单核细胞和粒细胞），我们在这里报告这种方法。我们还观察到，与先前提出的FFA多出口系统相比，基于尺寸的微珠分级的准确性显著提高。荧光显微镜说明了二维预聚焦的重要性，其中3-，7-和10-微米珠的样品混合物被分离成约束良好的颗粒流，并收集在各自的目标出口。淋巴细胞和粒细胞在其相应出口中的流式细胞术数据分别验证了同时分离的白细胞亚群具有高纯度（95.2 ± 0.6%和98.5 ± 0.7%）和高回收率（86.5 ± 10.9%和68.4 ± 10.6%）。在第三靶出口获得了相对低纯度和高回收率的单核细胞（25.2% ± 5.4%和83.1 ± 4.3%）。未观察到亚群偏倚。这些数据表明，在~100 μl/min的流速和~ 1 M细胞/ml的样本浓度下，白细胞亚群得到了前所未有的分离，这在声流系统中是以前没有报道过的。具有多个靶出口的二维预聚焦FFA是当前用于颗粒分级和细胞分离的方法的可行替代方案，需要最少的样品制备，并且降低分析时间和成本。
Microchip-based free flow acoustophoresis (FFA) in combination with two-dimensional cell prefocusing enables concurrent multiple target outlet fractionation of leukocytes into subpopulations (lymphocytes, monocytes and granulocytes); we report on this method here. We also observed significantly increased accuracy in size-based fractionation of microbeads as compared to previously presented FFA multiple outlet systems. Fluorescence microscopy illustrates the importance of two-dimensional prefocusing where a sample mixture of 3-, 7- and 10-micrometer beads are separated into well-confined particle streams and collected in their respective target outlets. Flow cytometry data for lymphocytes and granulocytes, respectively, in their corresponding outlets verify concurrent isolation of leukocyte subpopulations with high purity (95.2 ± 0.6% and 98.5 ± 0.7%) and high recovery (86.5 ± 10.9% and 68.4 ± 10.6%). A relatively low purity and high recovery of monocytes (25.2% ± 5.4% and 83.1 ± 4.3%) was obtained in the third target outlet. No subpopulation bias was observed. These data demonstrate an unprecedented separation of leukocyte subpopulations at flow rates of ~100 μl/min and ~1M cells/ml sample concentrations, not previously reported in acoustofluidic systems. Two-dimensional prefocusing FFA with multiple target outlets is a viable alternative to current methods for particle fractionation and cell isolation, requiring a minimum of sample preparation, and lowering analysis time and cost.