Improved quantitation of HER-2/neu gene copy number in breast tumor-derived DNA samples.
A Southern blot-based assay is presented that increases the simplicity and accuracy of the HER-2/neu gene copy number assignment in breast cancer DNA. Genomic DNA was hybridized simultaneously with probes corresponding to portions of HER-2/neu and a single-copy gene, myeloperoxidase. A unique restriction fragment was detected for each gene. The use of DNA probes of similar mass and equal specific activities resulted in a ratio of band intensities on the resultant autoradiograph that reported the ratio of gene copy numbers directly. Patient samples containing amplified levels of the HER-2/neu gene were identified by simple visual inspection of a single autoradiograph. Analysis of breast cancer samples alongside the cell line DNAs, representing a range of HER-2/neu gene copy numbers, permits visual quantitation of the tumors' gene copy numbers. The authors show that the HER-2/neu gene copy number can be determined accurately in marginally degraded DNA, a feature of some clinical samples.
Farkas
D H
DH
Department of Clinical Pathology, William Beaumont Hospital, Royal Oak, MI 48073-6769.
McMahon
L W
LW
Cai
P
P
Umek
R M
RM
D001345:Autoradiography; D014408:Biomarkers, Tumor; D015139:Blotting, Southern; D001943:Breast Neoplasms / Q000235:genetics*; D004273:DNA, Neoplasm / Q000235:genetics* / Q000378:metabolism; D003720:Densitometry; D003850:Deoxyribonuclease I; D066246:ErbB Receptors / Q000235:genetics*; D005784:Gene Amplification; D006801:Humans; D009195:Peroxidase / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D018719:Erb-b2 Receptor Tyrosine Kinases
D014408:Biomarkers, Tumor; D004273:DNA, Neoplasm; D003850:Deoxyribonuclease I; D066246:ErbB Receptors; D009195:Peroxidase; D011518:Proto-Oncogene Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases