Coates A, Tripp E
Sydney Melanoma Unit Laboratory, University of Sydney, Australia.
Melanoma Res. 1995 Apr;5(2):107-11. doi: 10.1097/00008390-199504000-00006.
Because the resistance of melanoma to cytotoxic therapy may be due to high intracellular glutathione (GSH), we wished to monitor GSH in melanoma cells during treatment. This required an assay suitable for very small samples. So we chose analytical flow cytometry. We calibrated the flow cytometric assays against biochemically determined GSH content in a range of cultured human melanoma cell lines, then applied the assays to fine-needle tumour biopsies. Mercury orange was the first fluorochrome suitable for use in a flow cytometer with a 488 nm light source, but many technical factors influenced the results. Chloromethyl fluorescein diacetate (CMFDA) allowed assay conditions less dependent on details of cell handling. Correlations between biochemical GSH content and fluorescence intensities in cell lines were good for mercury orange and CMFDA. CMFDA is the preferred fluorochrome for estimating cellular GSH content in biopsies from human melanomas. Tumours metastatic to or beyond regional lymph nodes had significantly more cells with high GSH than primary tumours or regional recurrences.
由于黑色素瘤对细胞毒性疗法产生抗性可能是由于细胞内谷胱甘肽（GSH）水平较高，我们希望在治疗过程中监测黑色素瘤细胞中的GSH含量。这需要一种适用于极少量样本的检测方法。因此，我们选择了分析流式细胞术。我们根据一系列培养的人黑色素瘤细胞系中通过生化方法测定的GSH含量，对流式细胞术检测进行了校准，然后将这些检测方法应用于细针肿瘤活检。汞橙是第一种适用于配备488 nm光源的流式细胞仪的荧光染料，但许多技术因素会影响检测结果。二乙酸羧甲基荧光素（CMFDA）使得检测条件较少依赖于细胞处理的细节。细胞系中生化GSH含量与荧光强度之间的相关性对于汞橙和CMFDA来说都很好。CMFDA是用于估计人黑色素瘤活检组织中细胞GSH含量的首选荧光染料。转移至区域淋巴结或超出区域淋巴结范围的肿瘤中，具有高GSH的细胞明显多于原发性肿瘤或区域复发病灶。
