Gupta Mamta, Naik Sita, Pandey C M, Dabadghao Sunil
Department of Immunology, Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, India.
Indian J Med Res. 2002 Jun;115:260-4.
BACKGROUND & OBJECTIVES: Drug sensitivity assays are useful in oncology practice for evaluating the sensitivity of malignant cells to anti-cancer drugs. The usefulness of such assays for the prediction of clinical response to therapy has also been demonstrated. The existing methods used for this purpose are time consuming and labour intensive. Here we report a simplified flow cytometry based assay for evaluating the in vitro drug sensitivity of leukaemic cells.
The chemo-sensitivity of three human leukaemic cell lines (a lymphoblastoid cell line, Jurkat; an erythroleukaemic cell line, K 562 and a myelomonocytic cell line HL-60) was investigated by flow cytometry. Flow cytometry was used to determine LD50 (50% inhibitory concentration) for prednisolone on Jurkat and daunorubicin on HL 60 and K 562 cell lines respectively. Per cent cell death could directly be assessed on a flow cytometer by measuring the fluorescence after staining with propidium iodide (PI). For comparison MTT assay was also performed using prednisolone on Jurkat and daunorubicin on HL-60.
Cytotoxic effect of drugs was found to be dose dependent. Mean LD50 of prednisolone for Jurkat cells by flow cytometry was 0.805 +/- 0.058 mg/ml and by MTT assay 0.866 +/- 0.115 mg/ml. Mean LD50 of daunorubicin for HL-60 was 1.96 +/- 0.05 micrograms/ml by flow cytometry and 1.90 +/- 0.282 micrograms/ml by MTT assay. The mean LD50 of daunorubicin to K 562 was 0.49 +/- 0.049 mg/ml by the flow cytometry method. The inter-assay variation for the LD50 by flow cytometry based assay was found to be 6, 14 and 10 per cent for Jurkat, HL-60 and K 562 respectively.
INTERPRETATION & CONCLUSION: We report a flow cytometry based drug-sensitivity assay for leukaemic cells, which uses a single dye staining and is rapid, technically simple and reproducible. The results compare well with the more commonly used MTT assay, which is labour intensive and time consuming. The limitation of our method is that it can only be used for studying cells in suspension and is therefore not suitable for adherent cell lines.
药物敏感性测定在肿瘤学实践中对于评估恶性细胞对抗癌药物的敏感性很有用。此类测定对预测治疗临床反应的有用性也已得到证实。用于此目的的现有方法既耗时又费力。在此，我们报告一种基于流式细胞术的简化测定方法，用于评估白血病细胞的体外药物敏感性。
通过流式细胞术研究了三种人类白血病细胞系（一种淋巴母细胞系，Jurkat；一种红白血病细胞系，K562；以及一种髓单核细胞系HL - 60）的化学敏感性。流式细胞术分别用于测定泼尼松龙对Jurkat细胞系以及柔红霉素对HL - 60和K562细胞系的半数抑制浓度（LD50）。通过用碘化丙啶（PI）染色后测量荧光，可在流式细胞仪上直接评估细胞死亡百分比。为作比较，还用泼尼松龙对Jurkat细胞系以及柔红霉素对HL - 60细胞系进行了MTT测定。
发现药物的细胞毒性作用呈剂量依赖性。通过流式细胞术测定，泼尼松龙对Jurkat细胞的平均LD50为0.805±0.058mg/ml，通过MTT测定为0.866±0.115mg/ml。通过流式细胞术测定，柔红霉素对HL - 60的平均LD50为1.96±0.05μg/ml，通过MTT测定为1.90±0.282μg/ml。通过流式细胞术方法，柔红霉素对K562的平均LD50为0.49±0.049mg/ml。基于流式细胞术的测定法对LD50的测定间变异在Jurkat、HL - 60和K562中分别为6%、14%和10%。
我们报告了一种用于白血病细胞的基于流式细胞术的药物敏感性测定法，该方法采用单一染料染色，快速、技术操作简单且可重复。结果与更常用的MTT测定法相比良好，MTT测定法既费力又耗时。我们方法的局限性在于它只能用于研究悬浮细胞，因此不适用于贴壁细胞系。
