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首页> 外文期刊>Cytometry: The Journal of the Society for Analytical Cytology >1-NGFR RECEPTOR IS A NEW FLOW CYTOMETRIC TOOL FOR RAPID CELL CYCLE-CORRELATED GENE THERAPY COMPLEMENTATION STUDIES IN VIABLE CELLS
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1-NGFR RECEPTOR IS A NEW FLOW CYTOMETRIC TOOL FOR RAPID CELL CYCLE-CORRELATED GENE THERAPY COMPLEMENTATION STUDIES IN VIABLE CELLS

机译:1-NGFR受体是用于活细胞快速细胞周期基因治疗互补研究的新型流式细胞仪

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Control of successful genetic complementation of cellular defects in heterogenous cell populations requires biochemical selection markers or cell analytical specifiers. With available gene therapy technologies, only a fraction of a cell population is transfected or transduced. We applied and optimized a novel dual-laser flow cytometric technique to analyze immediately the genetic complementation of cells dysregulated in cell proliferation and DNA repair. A novel bicistronic retrovirus carrying the normal Fanconi anemia gene (group C) and the cell surface marker gene 1-NGFR was constructed to analyze the normalization of the G2-phase cell cycle defect in DNA-repair-deficient FA(C) lymphoblastoid cells after transduction, Using a dual-laser multiparameter technology, we 1) analyzed the cell-cycle distribution of viable/dead cells using Hoechst 33342 (with ultraviolet Light), 2) identified the genetically complemented cells by FITC antibody labeling of the novel 1-NGFR surface marker (488 nn), and 3) recorded mitomycin C-induced cell death in nontransduced and transduced cells by propidium iodide, Artificial 1-NGFR expression is high and similar in ontogenetically and phylogenetically different cell populations. With this novel 1-NGFR marker technology, the success of gene therapy of cell-cycle dysregulation in even small subpopulations of cells can be recorded within 48 h, In addition to improved cell analysis, 1-NGFR surface marker expression can also be used for rapid generation of pure cell populations by column cell separation technologies. (C) 1997 Wiley-Liss, Inc. [References: 18]
机译:控制异种细胞群体中细胞缺陷的成功遗传互补需要生化选择标记或细胞分析说明物。利用可用的基因治疗技术,仅一部分细胞群体被转染或转导。我们应用并优化了一种新颖的双激光流式细胞仪技术,可以立即分析在细胞增殖和DNA修复中失调的细胞的遗传互补。构建了一种新型的具有正常范可尼贫血基因(C组)和细胞表面标记基因1-NGFR的双顺反子逆转录病毒,以分析DNA修复缺陷型FA(C)淋巴母细胞后G2期细胞周期缺陷的正常化转导,使用双激光多参数技术,我们1)使用Hoechst 33342(使用紫外线)分析了活/死细胞的细胞周期分布,2)通过FITC抗体标记的新型1-NGFR鉴定了遗传互补的细胞表面标记(488 nn)和3)记录了丝裂霉素C诱导的碘化丙啶在非转导和转导细胞中的细胞死亡,在个体发育和系统发育上不同的细胞群体中,人工1-NGFR表达很高且相似。借助这种新型的1-NGFR标记技术,即使在很小的细胞亚群中,细胞周期失调的基因治疗成功案例也可在48小时内记录下来。除了改进的细胞分析,1-NGFR表面标记的表达还可用于通过柱细胞分离技术快速生成纯细胞群体。 (C)1997 Wiley-Liss,Inc. [参考:18]

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