TY - JOUR
T1 - Clonal x-chromosome inactivation analysis of human tumors using a rapid, non-radioactive semi-quantitative pcr-based technique that allows analysis of fresh and archival material
AU - Kopp, Peter
AU - Totaler, Andreas
AU - Oeetreicher, Madeleins
AU - Jaggi, Rolf
AU - Borisch, Bettina
AU - Fey, Martin F.
AU - Larry Jameson, J.
PY - 1996/1/1
Y1 - 1996/1/1
N2 - The clonality of cell populations is important for the diagnosis and follow-up of disease. Most methods based on X-chromosome inactrvatkm require relatively large amounts of DNA. Two PCR-based strategies, the phosphoglycerate klnase (PGK) and the human androgen receptor (HUMARA) clonality assays, allow studies of small tissue samples. We made the HUMARA assay amenable to non-radioactive, semiquantitative analysis by performing Genescan analysis on an automated sequencer. The validity of the method was assessed by comparison with X-inactivation patterns obtained by Southern blot analysis with the probes M27B and PGK. 15 gastrointestinal carcinomas, 30 benign goiter nodules and normal peripheral leukocytes of 27 probands (12 4 15 years, 15 ï80 years) were analyzed. Furthermore, DNA extracted from formalin-fixed, paraffin-embedded tissue (FPT) was analyzed with the two PCR-based methods and compared with X-inactivation patterns generated with Southern analysis of high molecular weight (HMW) DNA. The Genescan-based HUMARA assay yields a high resolution of the short tandem repeats and direct semiquantitative resultsThe HUMARA assay is reliable in most patients; constitutive skewing in normal tissue (particularly in normal leukocytes) precludes clonal analysis in a few. Comparison of results obtained with HMW and FPT DNA yields consistent results with the HUMARA assay whereas the PGK PCR assay is unreliable. The HUMARA assay permits studies of selected areas of tissue sections without significant stromal components, thereby allowing correlation of histologie and genotype findings in archival material.
AB - The clonality of cell populations is important for the diagnosis and follow-up of disease. Most methods based on X-chromosome inactrvatkm require relatively large amounts of DNA. Two PCR-based strategies, the phosphoglycerate klnase (PGK) and the human androgen receptor (HUMARA) clonality assays, allow studies of small tissue samples. We made the HUMARA assay amenable to non-radioactive, semiquantitative analysis by performing Genescan analysis on an automated sequencer. The validity of the method was assessed by comparison with X-inactivation patterns obtained by Southern blot analysis with the probes M27B and PGK. 15 gastrointestinal carcinomas, 30 benign goiter nodules and normal peripheral leukocytes of 27 probands (12 4 15 years, 15 ï80 years) were analyzed. Furthermore, DNA extracted from formalin-fixed, paraffin-embedded tissue (FPT) was analyzed with the two PCR-based methods and compared with X-inactivation patterns generated with Southern analysis of high molecular weight (HMW) DNA. The Genescan-based HUMARA assay yields a high resolution of the short tandem repeats and direct semiquantitative resultsThe HUMARA assay is reliable in most patients; constitutive skewing in normal tissue (particularly in normal leukocytes) precludes clonal analysis in a few. Comparison of results obtained with HMW and FPT DNA yields consistent results with the HUMARA assay whereas the PGK PCR assay is unreliable. The HUMARA assay permits studies of selected areas of tissue sections without significant stromal components, thereby allowing correlation of histologie and genotype findings in archival material.
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M3 - Article
AN - SCOPUS:33749448877
VL - 44
JO - Journal of Investigative Medicine
JF - Journal of Investigative Medicine
SN - 1081-5589
IS - 3
ER -