Ishii M, Yamamuro T, Takeda T
Nihon Seikeigeka Gakkai Zasshi. 1982 Apr;56(4):293-304.
An attempt has been made to construct an assay of anti-cancer drug sensitivity which is suitable for use with primary culture cells from human bone and soft tissue tumors. The ability of cells to incorporate labeled nucleic acid precursors into acid precipitable material was assessed. Hela-S3 cells were employed to avoid inherited variability and heterogenity of primary cultures of human tumors. Labeled nucleic acid precursors were used not to assay the changes of DNA or RNA synthesis but to detect the viability of cells. A logarithm of counts per minute of incorporated labeled precursors is in proportion to a logarithm of viable cell numbers. This relationship was not influenced by the labeled precursor incubation time. Multiplate was used to provide large numbers of replicate cultures without the requirement of a large numbers of cells. Monolayer Hela-S3 cells which were seeded 24 hours earlier were incubated for three hours with various concentrations of drugs. After removal of drugs, cells were cultured for one week. In this period, Hela-S3 cells with no drug treatment became almost confluent and mitoses occurred about four times. Labeled precursors were incubated for three hours, and incorporated 3H-thymidine was counted. A standard curve of incorporated labeled precursor counts and viable cell numbers was drawn for every assay. The density inhibition of labeled nucleic acid precursor incorporation can be checked and connected with the standard curve, and viable cell numbers after drug exposure can be obtained from the standard curve. When percent survival is plotted against drug concentration, a sigmoid curve is obtained if the drug has dose dependent effect and then the 90% lethal dose (LD90) can be determined from the curve. LD90 was used as the index of anti-cancer drug sensitivity. If the drug has time dependent effect, percent survival of maximal inhibition was used as the index of drug sensitivity.
人们尝试构建一种抗癌药物敏感性检测方法，该方法适用于来自人骨和软组织肿瘤的原代培养细胞。评估了细胞将标记的核酸前体掺入酸不溶性物质中的能力。使用Hela-S3细胞以避免人肿瘤原代培养物的遗传变异性和异质性。使用标记的核酸前体不是为了检测DNA或RNA合成的变化，而是为了检测细胞的活力。掺入的标记前体的每分钟计数对数与活细胞数量的对数成比例。这种关系不受标记前体孵育时间的影响。使用多孔板提供大量重复培养物，而无需大量细胞。提前24小时接种的单层Hela-S3细胞与各种浓度的药物孵育3小时。去除药物后，细胞培养一周。在此期间，未接受药物处理的Hela-S3细胞几乎汇合，有丝分裂发生约四次。将标记前体孵育3小时，并对掺入的3H-胸腺嘧啶进行计数。每次检测绘制掺入的标记前体计数与活细胞数量的标准曲线。可以检查标记核酸前体掺入的密度抑制，并与标准曲线相关联，从标准曲线获得药物暴露后的活细胞数量。当将存活百分比与药物浓度作图时，如果药物具有剂量依赖性效应，则会得到一条S形曲线，然后可以从该曲线确定90%致死剂量（LD90）。LD90用作抗癌药物敏感性的指标。如果药物具有时间依赖性效应，则将最大抑制的存活百分比用作药物敏感性指标。
