Quantitative measurement of nuclear translocation events using similarity analysis of multispectral cellular images obtained in flow
Quantitative measurement of nuclear translocation events using similarity analysis of multispectral cellular images obtained in flow
复制标题
DOI：
10.1016/j.jim.2006.01.018
复制
发表时间：
2006-04-20
影响因子：
2.2
通讯作者：
Lynch, DH
中科院分区：
文献类型：
--
作者：
George, TC;Fanning, SL;Lynch, DH
NF-κ B的核转位启动许多基因的转录，其中许多基因对宿主防御至关重要。荧光素图像为基础的方法，量化这一事件历来利用贴壁细胞与大的细胞质-核面积比。然而，许多免疫相关细胞是天然非粘附的，并且具有小的胞质与核面积比。使用ImageStream（R）成像流式细胞仪，我们开发了一种新的方法，该方法使用来自每个细胞的核和NF-κ B图像的互相关分析来测量大群体中的核易位。该方法以剂量和时间依赖性方式准确测量具有小胞质面积的细胞中的NF-κ B易位。此外，NF-κ B易位在混合群体中包含的细胞亚群中被准确测量，并且该技术被成功地用于测量从人外周血获得的浆细胞样树突状细胞（PDC）中的IRF-7易位。本文描述的技术提供了一种客观和统计学上稳健的方法，用于测量具有特征性低胞质-核面积比的各种免疫相关细胞类型中的胞质-核分子易位事件。(c)2006 Elsevier B. V.保留所有权利。
Nuclear translocation of NF-kappa B initiates transcription of numerous genes, many of which are critical to host defense. Flourescent image-based methods that quantify this event have historically utilized adherent cells with large cytoplasm-to-nuclear area ratios. However, many immunologically relevant cells are naturally non-adherent and have small cytoplasm-to-nuclear area ratios. Using the ImageStream (R) imaging flow cytometer, we have developed a novel method that measures nuclear translocation in large populations using cross-correlation analysis of nuclear and NF-kappa B images from each cell. This approach accurately measures NF-kappa B translocation in cells with small cytoplasmic areas in dose- and time-dependent manners. Further, NF-kappa B translocation was accurately measured in a subset of cells contained in a mixed population and the technique was successfully employed to measure IRF-7 translocation in plasmacytoid dendritic cells (PDC) obtained from human peripheral blood. The techniques described here provide an objective and statistically robust method for measuring cytoplasmic to nuclear molecular translocation events in a variety of immunologically relevant cell types with characteristically low cytoplasm-to-nuclear area ratios. (c) 2006 Elsevier B.V. All rights reserved.