应用组织培养微量滴定试验检测天然产物中的细胞毒性剂。

Application of a tissue culture microtiter test for the detection of cytotoxic agents from natural products.

作者信息

Mirabelli C K, Bartus H, Bartus J O, Johnson R, Mong S M, Sung C P, Crooke S T

出版信息

J Antibiot (Tokyo). 1985 Jun;38(6):758-66. doi: 10.7164/antibiotics.38.758.

DOI:10.7164/antibiotics.38.758

PMID:4019319

Abstract

A method is described by which the growth inhibitory effects of cytotoxic compounds and fermentation broth cultures on adherent tumor cell lines can be quantitated. Cells are seeded into 96-well microtiter plates and 16 hours later the test compounds or broths are added to the wells. Cell growth is measured after three days (B16 mouse melanoma cells) or six days (HT-29, human colon carcinoma cells) by first fixing adherent cells, staining with Giemsa stain, washing away excess stain, then solubilizing stained cells with HCl. Absorbance is determined using a microELISA spectrophotometer and the data are transferred to and analyzed by a computer. The assay is rapid and reproducible and can be used to identify fermentation broths with cytotoxic components. Addition of DNA into the assay mixture (cells plus compound) inhibits the cytotoxic activities of certain DNA-reactive agents. The results of this study demonstrate the application of this assay system for primary and secondary evaluation of fermentation broths for in vitro antitumor activity.

摘要

本文描述了一种方法，通过该方法可以定量细胞毒性化合物和发酵液培养物对贴壁肿瘤细胞系的生长抑制作用。将细胞接种到96孔微量滴定板中，16小时后将测试化合物或发酵液加入孔中。三天后（B16小鼠黑色素瘤细胞）或六天后（HT-29人结肠癌细胞）测量细胞生长，方法是先固定贴壁细胞，用吉姆萨染色，洗去多余的染料，然后用盐酸溶解染色的细胞。使用微量ELISA分光光度计测定吸光度，并将数据传输到计算机并由计算机进行分析。该测定方法快速且可重复，可用于鉴定含有细胞毒性成分的发酵液。向测定混合物（细胞加化合物）中加入DNA可抑制某些DNA反应性试剂的细胞毒性活性。本研究结果证明了该测定系统在发酵液体外抗肿瘤活性的一级和二级评价中的应用。