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A simple G-banding technique following nonradioactive in situ hybridization
Abstracts
107
AN IMPROVED METHOD FOR OBTAINING A SIGNIFICANTLY LARGER NUMBER OF METAPHASES IN DISEASES OF THE BREAST. L.M.B.Cavali~ri*, S.R.Roaatto*, G.Vian-Borba**. * Departamento de Biologia Geral, CCB. ** Departamento de Patologia Aplicada, Legisla<;~io e Deontologia, HURNPr, Universidade Estadual de Londrina, PR, Brasil. Although breast tumors are very frequent, few cytogenetic data are available about them. One of the reasons for this lack of information is the difficulty in obtaining good quality cells for detailed chromosome analysis. Nine primary, untreated breast tumors (2 cystic hyperplasias, 2 fibroadenomas, 1 adenoma, and 4 carcinomas) were processed for tissue disaggregation using a 0.2% solution of collagenase IV (Sigma). The material was incubated in HAM F-10 medium (Sigma) supplemented with 20% fetal calf serum and antibiotics. When the cells were in exponential growth they were treated with trypsin-EDTA for 18 hours, after which fresh culture medium was used with 0.0016% colchicine added. After 18 hours, the material was processed by the standard technique for obtaining metaphase chromosomes. In all cases, the chromosomes obtained were elongated and of excellent morphology despite the long time of treatment with colchicine. The number of analyzable metaphases which were ideal for banding studies ranged from 23 to 140 on a slide from each case. Comparison of these results with those reported in the literature shows that this method results in a much larger number of metaphase cells which are also of markedly better quality for chromosome banding. CNPq, CAPES, CPG-UEL.
A SIMPLE G-BANDING TECHNIQUE FOLLOWING NONRADIOACTIVE IN SITU HYBRIDIZATION. Lisa M. Duncan (1), Erin Dietzsch (1), Christina Rudduck (1), 0.Margaret Garson(2) (1) Department of Medicine, University of Melbourne, Parkville, Victoria, Australia. (2) Department of Cytogenetics, St. Vincent's Hospital, Fitzroy, Victoria, Australia. A non-isotopic in situ hybridization (NISH) technique utilising a biotin-streptavidinpolyalkaline phosphatase complex has been found to be compatible with a simple G-banding protocol after detection of the hybridization signal. This method has been applied to analyze marker chromosomes in a cell line with a complex karyotype. The procedure requires no pre-treatment of cultures nor special optical equipment, and produces permanent preparations.
107
AN IMPROVED METHOD FOR OBTAINING A SIGNIFICANTLY LARGER NUMBER OF METAPHASES IN DISEASES OF THE BREAST. L.M.B.Cavali~ri*, S.R.Roaatto*, G.Vian-Borba**. * Departamento de Biologia Geral, CCB. ** Departamento de Patologia Aplicada, Legisla<;~io e Deontologia, HURNPr, Universidade Estadual de Londrina, PR, Brasil. Although breast tumors are very frequent, few cytogenetic data are available about them. One of the reasons for this lack of information is the difficulty in obtaining good quality cells for detailed chromosome analysis. Nine primary, untreated breast tumors (2 cystic hyperplasias, 2 fibroadenomas, 1 adenoma, and 4 carcinomas) were processed for tissue disaggregation using a 0.2% solution of collagenase IV (Sigma). The material was incubated in HAM F-10 medium (Sigma) supplemented with 20% fetal calf serum and antibiotics. When the cells were in exponential growth they were treated with trypsin-EDTA for 18 hours, after which fresh culture medium was used with 0.0016% colchicine added. After 18 hours, the material was processed by the standard technique for obtaining metaphase chromosomes. In all cases, the chromosomes obtained were elongated and of excellent morphology despite the long time of treatment with colchicine. The number of analyzable metaphases which were ideal for banding studies ranged from 23 to 140 on a slide from each case. Comparison of these results with those reported in the literature shows that this method results in a much larger number of metaphase cells which are also of markedly better quality for chromosome banding. CNPq, CAPES, CPG-UEL.
A SIMPLE G-BANDING TECHNIQUE FOLLOWING NONRADIOACTIVE IN SITU HYBRIDIZATION. Lisa M. Duncan (1), Erin Dietzsch (1), Christina Rudduck (1), 0.Margaret Garson(2) (1) Department of Medicine, University of Melbourne, Parkville, Victoria, Australia. (2) Department of Cytogenetics, St. Vincent's Hospital, Fitzroy, Victoria, Australia. A non-isotopic in situ hybridization (NISH) technique utilising a biotin-streptavidinpolyalkaline phosphatase complex has been found to be compatible with a simple G-banding protocol after detection of the hybridization signal. This method has been applied to analyze marker chromosomes in a cell line with a complex karyotype. The procedure requires no pre-treatment of cultures nor special optical equipment, and produces permanent preparations.