Maetzig Tobias, Morgan Michael, Schambach Axel
Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany; REBIRTH Cluster of Excellence, Hannover Medical School, Hannover, Germany.
Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany; REBIRTH Cluster of Excellence, Hannover Medical School, Hannover, Germany.
Exp Hematol. 2018 Nov;67:10-17. doi: 10.1016/j.exphem.2018.08.001. Epub 2018 Aug 8.
Hematopoiesis depends on the controlled differentiation of hematopoietic stem cells to mature cells with defined functions. Although each cell population within the hematopoietic hierarchy can be described by phenotypic markers, isolation of marker pure populations does not necessarily result in cells with homogeneous functionality. However, techniques that enable the efficient characterization of cell behavior with high resolution are limited. Although single-cell transplantation assays demand high mouse numbers and workload, sequencing-based fate tracking techniques require the destruction of the host cell, substantial financial resources, and bioinformatics expertise and suffer from a delay between sample acquisition and data interpretation. To make analyses more efficient, several laboratories recently developed flow cytometry-driven, fluorescence-based multiplexing approaches that enable parallel analysis of longitudinal behavior from multiple clonally derived cells or polyclonal populations. Although these fluorescent genetic barcoding systems are still in their infancy, their power lies in the use of retroviral vectors for gene marking of multiple populations with unique fluorescent color codes. Tracing of color-coded cells by flow cytometry guarantees the accessibility of information on population behavior in real time and at low cost, supports the prospective isolation of cells for downstream analyses, and can be applied to cell line models as well as to human- and animal-derived primary cells. Here, we discuss recent progress in the emerging field of fluorescent genetic barcoding for longitudinal multiplex cell tracking in biomedical research and how this technique will help to uncover mechanisms regulating cell behavior with clonal resolution in a reduced number of experimental samples.
造血作用依赖于造血干细胞向具有特定功能的成熟细胞的可控分化。尽管造血层级结构中的每个细胞群体都可以通过表型标记来描述，但分离标记纯合群体并不一定能得到功能均一的细胞。然而，能够以高分辨率有效表征细胞行为的技术有限。虽然单细胞移植试验需要大量小鼠且工作量大，但基于测序的命运追踪技术需要破坏宿主细胞、大量资金以及生物信息学专业知识，并且在样本采集和数据解读之间存在延迟。为了使分析更高效，几个实验室最近开发了基于流式细胞术的荧光多重分析方法，该方法能够对来自多个克隆衍生细胞或多克隆群体的纵向行为进行平行分析。尽管这些荧光遗传条形码系统仍处于起步阶段，但其优势在于使用逆转录病毒载体对多个群体进行基因标记，赋予独特的荧光颜色编码。通过流式细胞术追踪颜色编码的细胞可确保实时、低成本地获取群体行为信息，支持对细胞进行前瞻性分离以进行下游分析，并且可应用于细胞系模型以及人源和动物源原代细胞。在这里，我们讨论了生物医学研究中用于纵向多重细胞追踪的荧光遗传条形码这一新兴领域的最新进展，以及该技术将如何有助于在减少实验样本数量的情况下以克隆分辨率揭示调节细胞行为的机制。
