Flow cytometric analysis of DNA and nuclear protein in paraffin-embedded tissue.
Flow cytometric analysis of DNA and nuclear protein in paraffin-embedded tissue.
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石蜡包埋组织中 DNA 和核蛋白的流式细胞术分析。
DOI：
10.1002/cyto.990140213
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Block,NL
中科院分区：
文献类型：
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作者：
Ciancio,G;Pollack,A;Block,NL
以前，我们描述了同时定量的DNA和核蛋白在未固定的组织从实体瘤。由此产生的2参数流式细胞术分析与单独的DNA分析相比具有多个优势。在本报告中，我们描述了福尔马林固定石蜡包埋组织分析技术的改进。通过水合切片、在0.5%胃蛋白酶溶液中孵育、洗涤并重悬于含非离子去污剂的缓冲液中制备石蜡包埋材料。然后在核糖核酸酶存在下用异硫氰酸荧光素和碘化丙啶对细胞核进行染色。已经分析了几种实体瘤组织类型，包括乳腺、结肠、肾脏和胸腺。当胃蛋白酶初始处理时间为1.5 h而不是0.5 h时，获得最佳结果。胃蛋白酶处理1.5小时改善了DNA和核蛋白参数的CV，并且似乎没有降低核蛋白水平或引起核的显著崩解。未固定和固定石蜡包埋组织的DNA/核蛋白直方图相似。由于肿瘤细胞核通常比DNA二倍体细胞核具有更高的蛋白质水平，因此该技术减少了群体重叠，并允许较少的DNA非整倍性的主观鉴定。© 1993 Wiley利斯公司
Previously we described the simultaneous quantification of DNA and nuclear protein in unfixed tissue from solid tumors. The resultant 2 parameter flow cytometric analysis has several advantages over that of DNA alone. In this report, we describe a modification of the technique for the analysis of formalin‐fixed paraffin‐embedded tissue. Paraffin‐embedded material was prepared by hydrating sections, incubating in 0.5% pepsin solution, washing, and resuspending in buffer containing nonionic detergent. The nuclei were then stained with fluorescein isothiocyanate and propidium iodide in the presence of ribonuclease. Several solid tumor tissue types have been analyzed, including breast, colon, kidney, and thymus. The best results were obtained when the initial pepsin treatment was for 1.5 h, instead of 0.5 h. Pepsin treatment for 1.5 h improved the CVs of both the DNA and nuclear protein parameters, and did not appear to reduce nuclear protein levels or to cause significant disintegration of nuclei. The DNA/nuclear protein histograms of unfixed and fixed, paraffin‐embedded tissue were similar. Since tumor nuclei typically have higher protein levels than DNA‐diploid nuclei, the technique reduces population overlapping and permits less subjective identification of DNA aneuploidy. © 1993 Wiley‐Liss, Inc.