摘要
肿瘤细胞由多聚甲醛直接固定在硝酸纤维素膜上,经蛋白酶消化后与地高辛配基(di-goxigenin)标记的癌基因探针进行DNA-RNA杂交,洗膜后用酶联免疫法显色,可以半定量的确定癌基因在肿瘤细胞中的表达状况。该方法不需提取RNA,因此操作简便易行。由于利用地高辛配基标记探针,克服了同位素放射性和蜕变带来的诸多不便。实验将此法用于肺癌细胞经诱导分化后癌基因表达改变以及白血病细胞中癌基因表达情况研究,5×10~3个细胞即可获得阳性杂交信号。
A simple and sensitive method for detecting oncogene mRNA is performed by fixing tumor cells directly onto nitrocellulose paper, followed by in situ RNA hybridization with digoxigenin-labelled oncogene probes. The human lung large cell carcinoma cell line PLA801-D95(before and after induction of differentiation with retinoid acid) and leukemic cells from clinical patients were used as target cells. The results showed that 5000 cells were sufficient to give a positive signal for oncogne mRNA. Compared with RNA dot blot, it was found that this technique was quick, simple, sensitive and reproducible. It can be used to study gene expression in tumor cells as well as other cells.
出处
《中国医学科学院学报》
CAS
CSCD
北大核心
1991年第6期439-442,共4页
Acta Academiae Medicinae Sinicae
关键词
核酸杂交
地高辛配基
癌基因
DNA-RNA hybridization digoxigenin oncogene
