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Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.

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...

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Autores principales: Abeln, E. C., Corver, W. E., Kuipers-Dijkshoorn, N. J., Fleuren, G. J., Cornelisse, C. J.
Formato: Texto
Lenguaje:English
Publicado: Nature Publishing Group 1994
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2033519/
https://www.ncbi.nlm.nih.gov/pubmed/8054273
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author Abeln, E. C.
Corver, W. E.
Kuipers-Dijkshoorn, N. J.
Fleuren, G. J.
Cornelisse, C. J.
author_facet Abeln, E. C.
Corver, W. E.
Kuipers-Dijkshoorn, N. J.
Fleuren, G. J.
Cornelisse, C. J.
author_sort Abeln, E. C.
collection PubMed
description 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. IMAGES:
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spelling pubmed-20335192009-09-10 Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity. Abeln, E. C. Corver, W. E. Kuipers-Dijkshoorn, N. J. Fleuren, G. J. Cornelisse, C. J. Br J Cancer Research Article 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. IMAGES: Nature Publishing Group 1994-08 /pmc/articles/PMC2033519/ /pubmed/8054273 Text en https://creativecommons.org/licenses/by/4.0/This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit https://creativecommons.org/licenses/by/4.0/.
spellingShingle Research Article
Abeln, E. C.
Corver, W. E.
Kuipers-Dijkshoorn, N. J.
Fleuren, G. J.
Cornelisse, C. J.
Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
title Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
title_full Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
title_fullStr Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
title_full_unstemmed Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
title_short Molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
title_sort molecular genetic analysis of flow-sorted ovarian tumour cells: improved detection of loss of heterozygosity.
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2033519/
https://www.ncbi.nlm.nih.gov/pubmed/8054273
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