A Versatile Platform for Comprehensive Chip-Based Explorative Cytometry
A Versatile Platform for Comprehensive Chip-Based Explorative Cytometry
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DOI：
10.1002/cyto.a.20668
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发表时间：
2009-04-01
期刊：
影响因子：
3.7
通讯作者：
Hansen, Gesine
中科院分区：
文献类型：
--
作者：
Hennig, Christian;Adams, Nico;Hansen, Gesine
免疫系统的巨大复杂性的分析越来越受到当前方法的技术限制的阻碍，特别是对于含有少量细胞的样品的多参数和功能分析。我们在这里提出了一种方法，这是基于逐步的功能操作和分析的活的免疫细胞，是自我固定在微流控芯片内，使用自动落射荧光显微镜克服目前的限制，全面的免疫表型。与流式细胞术交叉验证显示10倍增加的灵敏度和可比的特异性。通过使用小的样品体积和细胞数量（2-10 μ l，低至20，000个细胞），我们能够分析几乎无限数量的细胞内和表面标记物，甚至在活的免疫细胞上。我们通过（1）稀有细胞的鉴定和表型分析，（2）非常有限的样本量的综合分析，以及（3）体外分化后人类B细胞的深度免疫表型分析，证实了这种方法的科学和诊断潜力。最后，我们提出了一个信息模型的注释和比较的细胞计数数据，通过使用基于本体的方法。本文介绍的基于芯片的细胞仪被证明是一种非常有用的工具，可以逐步探索具有几乎无限数量的表面和细胞内标记物的珍贵的含小细胞样品。(C)2008年国际细胞计数促进学会
Analysis of the immense complexity of the immune system is increasingly hampered by technical limitations of current methodologies, especially for multiparameter- and functional analysis of samples containing small numbers of cells. We here present a method, which is based on the stepwise functional manipulation and analysis of living immune cells that are self-immobilized within microfluidic chips using automated epi-fluorescence microscopy overcoming current limitations for comprehensive immunophenotyping. Crossvalidation with flow cytometry revealed a 10-fold increased sensitivity and a comparable specificity. By using small sample volumes and cell numbers (2-10 mu l, down to 20,000 cells), we were able to analyze a virtually unlimited number of intracellular and surface markers even on living immune cells. We exemplify the scientific and diagnostic potential of this method by (1) identification and phenotyping of rare cells, (2) comprehensive analysis of very limited sample volume, and (3) deep immunophenotyping of human B-cells after in vitro differentiation. Finally, we propose an informatic model for annotation and comparison of cytometric data by using an ontology-based approach. The chip-based cytometry introduced here turned out to be a very useful tool to enable a stepwise exploration of precious, small cell-containing samples with an virtually unlimited number of surface- and intracellular markers. (C) 2008 International Society for Advancement of Cytometry