Confocal assay for invasion: Use of propidium iodide fluorescence and laser reflectance to quantify the rate of migration of cells through a matrix
Confocal assay for invasion: Use of propidium iodide fluorescence and laser reflectance to quantify the rate of migration of cells through a matrix
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DOI：
10.1002/1097-0320(20000801)40:4
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发表时间：
2000-08-01
期刊：
影响因子：
--
通讯作者：
Givan, AL
中科院分区：
文献类型：
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作者：
Benbow, U;Orndorff, KA;Givan, AL
背景：大多数用于测量侵袭的测定是基于对已经完全迁移通过的细胞的数目的手动计数。商用涂层过滤器。我们在这里描述了一种共聚焦荧光成像方法，它可以评估细胞侵入matrix.Methods的相对速率：将细胞接种在基质上并孵育一段时间后，将细胞固定并用RNase处理。然后加入碘化丙啶以染色双链DNA。共聚焦显微镜系统用于获得高分辨率图像的红色碘化丙啶荧光和激光反射率从光学部分在矩阵中的深度增加。具有高激光反射率的部分标记基质的顶部。结果：数据计算为每个部分中高于背景的红色荧光的总面积，并绘制为所有部分中总荧光面积的百分比。因为核进入基质的距离可以通过从基质的反射上表面测量来计算，该方法可用于评估细胞迁移速率和比较不同细胞在不同条件下侵入不同基质的能力。(C)2000 Wiley-Liss，Inc.
Background: Most assays used to measure invasion are based on manual counting of the number of cells that have migrated completely through. commercial coated filters. We describe here a confocal fluorescence-imaging method chat can assess the relative rates of invasion of cells into a matrix.Methods: After being seeded on the matrix and a period of incubation, the cells are fixed and treated with RNase. Propidium iodide is then added to stain the double-stranded DNA. A confocal microscope system is used to obtain high-resolution images of the red propidium iodide fluorescence and laser reflectance from optical sections at increasing depths in the matrix. The section with high laser reflectance marks the top of the matrix.Results: Data were calculated as the total area of red fluorescence above background in each section and were plotted as a percentage of the summed fluorescent areas in all sections.Conclusions: Because the distance into the matrix of the nuclei can be calculated by measuring from the reflective upper surface of the matrix, the method is useful for assessing the rate of cell migration and for comparing the ability of different cells to invade through different matrices under varying conditions. (C) 2000 Wiley-Liss, Inc.