A reliable tool to determine cell viability in complex 3-D culture: The acid phosphatase assay
A reliable tool to determine cell viability in complex 3-D culture: The acid phosphatase assay
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DOI：
10.1177/1087057107306839
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发表时间：
2007-10-01
影响因子：
--
通讯作者：
Kunz-Schughart, Leoni A.
中科院分区：
文献类型：
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作者：
Friedrich, Juergen;Eder, Wolfgang;Kunz-Schughart, Leoni A.
基于细胞的分析比无细胞测试系统更复杂，但仍然反映了高度人工的细胞环境。将器官类型三维(3-D)培养系统纳入主流药物开发过程的讨论越来越多，但受到复杂的方法学要求的严重限制。这项研究的目的是探索一组标准的分析方法，为基于多细胞肿瘤球体的抗肿瘤药物试验中细胞存活的快速常规分析提供一种易于操作的标准化方案。对2个结肠癌细胞系的球体进行了鉴定。其中一个测试系统可以可靠地用于确定球体中的细胞活性。作者验证了酸性磷酸酶测定法(APH)适用于96孔板中的单个球体，不需要球体解离，对直径分别为4万个细胞和8万个细胞的HT29和HCT-116球体具有线性和高灵敏度。用5-氟尿嘧啶、伊立替康和C-1311处理HT29和HCT-116细胞显示，与单层培养相比，3D培养的药物效率严重降低，这一点根据文献数据进行了讨论。本文提出的实验方案是在抗肿瘤药物筛选方案中建立复杂的3-D体外系统的一个小但实质性的贡献。
Cell-based assays are more complex than cell-free test systems but still reflect a highly artificial cellular environment. Incorporation of organotypic 3-dimensional (3-D) culture systems into mainstream drug development processes is increasingly discussed but severely limited by complex methodological requirements. The objective of this study was to explore a panel of standard assays to provide an easy-handling, standardized protocol for rapid routine analysis of cell survival in multicellular tumor spheroid-based antitumor drug testing. Spheroids of 2 colon carcinoma cell lines were characterized for evaluation. One of the assay systems tested could reliably be used to determine cell viability in spheroids. The authors verified that the acid phosphatase assay (APH) is applicable for single spheroids in 96-well plates, does not require spheroid dissociation, and is linear and highly sensitive for HT29 and HCT-116 spheroids up to diameters of 650 mu m and 900 gm, consisting of 40,000 and 80,000 cells, respectively. Treatment of HT29 and HCT-116 cells with 5-fluorouracil, Irinotecan, and C-1311 revealed critically reduced drug efficacies in 3-D versus monolayer culture, which is discussed in light of literature data. The experimental protocol presented herein is a small but substantial contribution to the establishment of sophisticated 3-D in vitro systems in the antitumor drug screening scenario.