Minimal illegitimate levels of cytokeratin K19 expression in mononucleated blood cells detected by a reverse transcription PCR method (RT-PCR)
| Autor: | P. Bolufer-Gilabert, A. Sempere-Talens, J Palau-Pérez, E. Barragán-González, J. De la Rubia-Comos, S Bonanad-Boix, M. Sanz-Alonso, José Antonio López-Guerrero |
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| Rok vydání: | 1997 |
| Předmět: |
DNA
Complementary Serial dilution Clinical Biochemistry Gene Expression Breast Neoplasms Biology Biochemistry Peripheral blood mononuclear cell Polymerase Chain Reaction Sensitivity and Specificity Complementary DNA Gene expression Humans Leukemia Biochemistry (medical) RNA General Medicine DNA Neoplasm Mononucleated Blood Cell Hematopoietic Stem Cells Molecular biology Hodgkin Disease Reverse transcription polymerase chain reaction Real-time polymerase chain reaction Evaluation Studies as Topic Case-Control Studies Leukocytes Mononuclear Keratins Female |
| Zdroj: | CLINICA CHIMICA ACTA r-IIS La Fe. Repositorio Institucional de Producción Científica del Instituto de Investigación Sanitaria La Fe instname |
| ISSN: | 0009-8981 |
| Popis: | Cytokeratin K19 (CK19) expression was evaluated by a reverse transcription PCR method (RT-PCR) in the RNA obtained from peripheral blood stem cell collections (PBSC) from four patients with breast cancers (BC) and 34 mononucleated blood cell (MBC) negative controls (17 PBMC from normal subjects 12 PBSC from different types of leukaemias--M3, M4Eo, M2, etc.--and two from patients with Hodgkin's lymphoma; and three bone marrow (BM) collections). Two BC tissues were taken as positive controls. The method studied (Datta YH, Paul T, Adams PT, Drobyski WR. Sensitive detection of occult breast cancer by reverse transcription polymerase chain reaction. J Oncol 1994;12:475-8) is sensitive enough to allow the detection of CK19 transcripts in a 10(-6) dilution of cDNA reverse transcribed from 1 microgram of BC RNA, but CK19 transcripts were also detected in 64% of the RNA obtained from the MBC controls. However, the amplified product detected in the control samples represents the transcript of the CK19 gene as confirmed by the results of Mae III digestion. It should be pointed out that although the CK19 expression was detected, the levels of expression in PBMC were almost negligible for they disappeared at 1:5 cDNA dilution. Moreover, a direct relationship between the number of BC cells added to PBMC and the increasing dilution levels of the cDNA necessary to prevent CK19 expression was observed. This allows us to conclude that the cDNA dilutions make it possible to distinguish the false from the true positive samples and that, in addition, the cDNA dilutions inform about the degree of BC cell contamination. |
| Databáze: | OpenAIRE |
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