Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
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DOI：
10.1038/bjc.1994.289
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发表时间：
1994-08
影响因子：
8.8
通讯作者：
Cornelisse, C. J.
中科院分区：
文献类型：
--
作者：
Abeln, E. C. A.;Cover, W. E.;Kuipers-Dijkshoorn, N. J.;Fleuren, G.-J.;Cornelisse, C. J.
杂合性缺失（洛）的检测通常在均质化的肿瘤标本上进行。在这种类型的分析中，必须排除具有低百分比肿瘤细胞的样品，并且可能的肿瘤内异质性被掩盖。在这项研究中，我们报告了聚合酶链反应（PCR）驱动的洛杂合性缺失检测的应用，共使用染色体1 q、3 p、3 q、4 p、6 p、6 q、11 p、11 q、17 p、17 q、18 p、18 q、Xp和Xq的22个微卫星标记对来自新鲜和石蜡包埋的卵巢肿瘤组织的流式细胞进行检测。滴定实验表明，洛缺失可以用低至100个细胞当量的DNA检测。明确的洛的例子可以检测到在分选的非整倍体部分从一个单侧和两个双侧卵巢肿瘤从三个病人。在两个样品中，分选部分小于总样品的10%。来自同一患者的双侧肿瘤显示一个标志物（病例OV 64）和两个标志物（病例OV 69）的相同等位基因的丢失，指示它们的单克隆起源。使用两种不同的卵巢肿瘤标志物（MOv 18和BMA 180），抗细胞角蛋白单克隆抗体（MAb）（M9），抗波形蛋白单克隆抗体（V9）和抗泛上皮抗原17-1A的单克隆抗体对第四卵巢癌患者的新鲜腹水细胞的多参数流式细胞术被用来研究可能的肿瘤内异质性。我们发现存在至少三个表型不同的人群，其中二倍体，角蛋白阳性，波形蛋白阴性的人口表现出类似的洛合性缺失模式的非整倍体人口（DNA指数= 1.7），表明其肿瘤起源。同样的洛缺失模式也出现在该患者的网膜转移瘤中，其非整倍体DNA指数也为1.7。二倍体和非整倍体肿瘤细胞群体具有相同的洛缺失模式，这表明观察到的等位基因缺失事件发生在非整倍体发生之前。因此，流式细胞PCR是研究肿瘤克隆多样性的重要工具。
Detection of loss of heterozygosity (LOH) is usually performed on homogenised tumour specimens. In this type of analysis samples with a low percentage of tumour cells have to be excluded and possible intra-tumour heterogeneity is obscured. In this study we report the application of polymerase chain reaction (PCR)-driven LOH detection with in total 22 microsatellite markers for chromosome 1q, 3p, 3q, 4p, 6p, 6q, 11p, 11q, 17p, 17q, 18p, 18q, Xp and Xq on flow-sorted cells from fresh and paraffin-embedded ovarian tumour tissue. Titration experiments showed that LOH can be detected with as few as 100 cell equivalents of DNA. Clear examples of LOH could be detected in the sorted aneuploid fractions from one unilateral and two bilateral ovarian tumours from three patients. In two samples the sorted fraction was less than 10% of the total sample. The bilateral tumours from the same patient showed loss of identical alleles for one marker (case OV64) and two markers (case OV69), indicative of their monoclonal origin. Multiparameter flow cytometry using two different ovarian tumour markers (MOv18 and BMA180), an anti-cytokeratin monoclonal antibody (MAb) (M9), an anti-vimentin MAb (V9) and a MAb against the panepithelial antigen 17-1A on the fresh ascites cells of the fourth ovarian cancer patient was used to investigate possible intra-tumour heterogeneity. We showed the presence of at least three phenotypically different populations, of which the diploid, keratin-positive, vimentin-negative population showed a similar LOH pattern as the aneuploid population (DNA index = 1.7), indicative of its neoplastic origin. The same LOH pattern was shown in an omentum metastasis from this patient also having the same aneuploid DNA index of 1.7. The sharing of the same LOH pattern by the diploid and aneuploid tumour cell populations suggests that the observed allele loss events occurred before the development of aneuploidy. PCR on flow-sorted cells is thus an important tool to study clonal diversity in tumours.
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影响因子：
8.8
作者：
Foulkes, W D;Ragoussis, J;Stamp, G W;Allan, G J;Trowsdale, J
通讯作者：
Trowsdale, J
DOI：
10.1073/pnas.77.8.4914
发表时间：
1980-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
HOFFMAN, RA;KUNG, PC;GOLDSTEIN, G
通讯作者：
GOLDSTEIN, G
影响因子：
4.4
作者：
DEVILEE, P;VANDENBROEK, M;CORNELISSE, CJ
通讯作者：
CORNELISSE, CJ
影响因子：
8.8
作者：
Foulkes, W D;Campbell, I G;Stamp, G W;Trowsdale, J
通讯作者：
Trowsdale, J
影响因子：
6.4
作者：
PHILLIPS, N;ZIEGLER, M;XYNOS, F
通讯作者：
XYNOS, F