Sensitive determination of cell number using the CyQUANT® cell proliferation assay
Sensitive determination of cell number using the CyQUANT® cell proliferation assay
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DOI：
10.1016/s0022-1759(01)00404-5
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发表时间：
2001-08-01
影响因子：
2.2
通讯作者：
Singer, VL
中科院分区：
文献类型：
--
作者：
Jones, LJ;Gray, M;Singer, VL
我们在这里描述的CyQUANT（（R））细胞增殖试验，一个高灵敏度，基于荧光的微孔板测定法，用于确定培养细胞的数量的发展和表征。该测定采用CyQUANT GR染料，其在与细胞核酸结合时产生大的荧光增强，其可以使用标准荧光素激发和发射波长来测量。染料-核酸复合物的荧光发射与使用多种细胞类型的大范围内的细胞数量线性相关。在推荐的测定条件下，标准曲线呈线性（r（2）> 0.995），根据细胞类型，使用单一染料浓度可检测少至10-50个细胞，多至25，000 - 50，000个细胞。增加染料浓度将测定的线性范围扩展至100，000 - 250，000个细胞。使用该测定法进行的细胞增殖和生长抑制研究的结果与使用其他标准测定法进行的已发表研究中获得的结果相似。血清刺激的细胞增殖的CyQUANT测定法测量值与使用[H-3]-胸苷进行的测量值相关性良好。Also.该测定用于分析细胞DNA或RNA含量。在该方案中增加了核酸酶消化步骤，该测定方法简单方便，没有洗涤步骤，并且易于自动化。(C)2001 Elsevier Science B. V.保留所有权利。
We describe here the development and characterization of the CyQUANT((R)) cell proliferation assay, a highly sensitive, fluorescence-based microplate assay for determining numbers of cultured cells. The assay employs CyQUANT GR dye, which produces a large fluorescence enhancement upon binding to cellular nucleic acids that can be measured using standard fluorescein excitation and emission wavelengths. The fluorescence emission of the dye-nucleic acid complexes correlated linearly with cell number over a large range using a wide variety of cell types. Under the recommended assay conditions, standard curves were linear(r(2) > 0.995), detecting as few as 10-50 cells and as many as 25,000-50,000 cells with a single dye concentration, depending on cell type. Increasing the dye concentration extended the linear range of the assay to 100,000-250,000 cells. Results of cell proliferation and growth inhibition studies with the assay were similar to those obtained in published studies using other standard assays. CyQUANT assay measurements of serum-stimulated cell proliferation correlated well with measurements made using [H-3]-thymidine. Also. the assay was used to analyze cellular DNA or RNA content. with the addition of a nuclease digestion step to the protocol, The assay procedure is simple and convenient, with no wash steps, and is readily amenable to automation. (C) 2001 Elsevier Science B.V. All rights reserved.