Development of a mast cell-based biosensor
Development of a mast cell-based biosensor
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DOI：
10.1016/j.bios.2007.10.007
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发表时间：
2008-02-28
影响因子：
12.6
通讯作者：
Holowka, David
中科院分区：
文献类型：
--
作者：
Curtis, Theresa;Naal, Rose Mary Z. G.;Holowka, David
一种基于肥大细胞的生物传感器已经开发出来，可以将这些细胞用于多种应用，包括药物筛选、环境监测、临床诊断和国土安全。大鼠嗜碱性白血病（RBL）肥大细胞为生物传感器的应用提供了极好的潜力，因为它们是健壮的，并且在抗原添加后几分钟内就会发生剧烈的胞外反应。为了监测肥大细胞的活化，荧光染料被加载到细胞中，作为分泌颗粒碱化、钙通量或活性氧生成的指标。这些荧光分析有效地测量活化抗原刺激的RBL肥大细胞，检测抗原的皮摩尔灵敏度。为了证明这种基于肥大细胞的生物传感器在检测微生物病原体方面的实用性，通过将IgE抗体的Fc区与CD14（一种脂多糖受体）融合，制备了IgE嵌合蛋白。这种嵌合蛋白有能力与大肠杆菌和单核增生李斯特菌结合，也能与肥大细胞上的IgE受体结合，从而刺激对细菌的信号反应。用钙指示剂Fluo-4标记的RBL肥大细胞被证明对大肠杆菌有反应，只有当用嵌合蛋白致敏时，从而证明了一种高度通用的细菌污染生物传感器。(C) 2007 Elsevier B.V.版权所有
A mast cell-based biosensor has been developed to enable the use of these cells in numerous applications including pharmaceutical screening, environmental monitoring, clinical diagnosis and homeland security. Rat basophilic leukemia (RBL) mast cells offer excellent potential for biosensor applications because they are robust and undergo a dramatic exocytotic response within minutes of antigen addition. To monitor mast cell activation, fluorescent dyes were loaded into the cells and used as indicators of alkalinization of secretory granules, calcium fluxes or generation of reactive oxygen species. These fluorescence assays efficiently measure activation of antigen-stimulated RBL mast cells, detecting the antigen with picomolar sensitivity. To demonstrate the utility of this mast cell-based biosensor for detection of microbial pathogens, an IgE chimeric protein was created by fusing the Fc region of the IgE antibody to CD14, a receptor for lipopolysaccharide. This chimeric protein has the capacity to bind to Escherichia coli and Listeria monocytogenes and also to IgE receptors on the mast cells, thereby stimulating a signaling response to bacteria. RBL mast cells labeled with the calcium indicator Fluo-4 are shown to be responsive to E. coli, only when sensitized with the chimeric protein, thus demonstrating a highly versatile biosensor for bacterial contamination. (C) 2007 Elsevier B.V. All rights reserved.