title: A new nested primer pair improves the specificity of CK-19 mRNA detection by RT-PCR in occult breast cancer cells.
abstract: Reverse transcription polymerase chain reaction (RT-PCR) of cytokeratin-19 (CK-19) has been widely used to detect small numbers of circulating malignant epithelial cells in the bone marrow or the peripheral blood of patients with breast cancer. However, a high percentage of false positive results has been recorded and conflicting reports question the clinical relevance of this technical approach. We demonstrate that the use of a new nested primer pair for CK-19 RT-PCR avoids false positive results without affecting the sensitivity of the assay. Our experiments were carried out using MCF-7 cells alone or mixed with peripheral blood mononuclear cells (PBMNC) of healthy donors. The results were also validated in a large series of healthy donors and in a preliminary study on a limited number of patients with breast cancer, thus suggesting that our assay is feasible for application in the clinical evaluation of occult malignant epithelial cells.
authors.affiliation: Department of Oncology and Hemato-Oncology, P.F. Calvi Hospital, Noale, Venice, Italy.
authors.forename: E
authors.initials: E
authors.lastname: Balducci
authors.forename: G
authors.initials: G
authors.lastname: Azzarello
authors.forename: L
authors.initials: L
authors.lastname: Valori
authors.forename: L
authors.initials: L
authors.lastname: Toffolatti
authors.forename: L
authors.initials: L
authors.lastname: Bolgan
authors.forename: M T
authors.initials: MT
authors.lastname: Valenti
authors.forename: M
authors.initials: M
authors.lastname: Bari
authors.forename: G L
authors.initials: GL
authors.lastname: Pappagallo
authors.forename: S
authors.initials: S
authors.lastname: Ausoni
authors.forename: O
authors.initials: O
authors.lastname: Vinante
mesh_terms: D000328:Adult; D000368:Aged; D001483:Base Sequence; D014408:Biomarkers, Tumor / Q000235:genetics; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology*; D045744:Cell Line, Tumor; D017931:DNA Primers / Q000235:genetics*; D006801:Humans; D007633:Keratins / Q000235:genetics*; D008875:Middle Aged; D008969:Molecular Sequence Data; D009382:Neoplasms, Unknown Primary / Q000235:genetics* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*
chemical_list: D014408:Biomarkers, Tumor; D017931:DNA Primers; D012333:RNA, Messenger; D007633:Keratins