Comparison of the relative expression of exon 3 deleted estrogenreceptor (ER) variant (ERD3) messenger RNA between breast tumor and adjacentmatched normal breast samples. (A) Total RNA extracted from frozentissue sections from tumor (T) and adjacent normal (N) breast tissue sampleswas reverse transcribed and polymerase chain reaction (PCR) amplified using D5Uand D5L primers. Radioactive PCR products were separated on a 6% acrylamide geland visualized by autoradiography. Bands that migrated at 483 and 344 basepairs were identified as corresponding to wild-type (WT)-ER and exon 5 deletedER variant (ERD5) messenger RNA, respectively. C, negative control (nocomplementary DNA added during the PCR reaction). (B) For each case,signals corresponding to ERD5 variant messenger RNA were quantified andexpressed in arbitrary units for tumor (black column) and normal (white column)components. For each sample, the mean and the standard deviation of at leastthree different PCR assays are indicated. Cases are sorted by ER status (blackbottom lane) and progesterone receptor (PR) status (gray bottom lane). Samplesthat failed to have three measurable signals in the four experiments performedin both normal and neoplastic components were not included in the statisticalanalysis. The significance of the differences between tumor and normal matchedcomponents within each subgroup, as tested using the Wilcoxon matched-pairtest, is indicated where P <0.05. m, molecular weight marker.