title: A quantitative reverse transcriptase polymerase chain reaction-based assay to detect carcinoma cells in peripheral blood.
abstract: The presence of tumour cells in the circulation may predict disease recurrence and metastasis. To improve on existing methods of cytological or immunocytological detection, we have developed a sensitive and quantitative technique for the detection of carcinoma cells in blood, using the reverse transcriptase polymerase chain reaction (RT-PCR) identifying transcripts of the pancarcinoma-associated tumour marker EGP-2 (KSA or 17-1A antigen). The amount of EGP2 mRNA was quantified using an internal recombinant competitor RNA standard with known concentration and which is both reversely transcribed and co-amplified in the same reaction, allowing for a reliable assessment of the initial amount of EGP2 mRNA in the sample. Calibration studies, seeding blood with MCF-7 breast carcinoma cells, showed that the assay can detect ten tumour cells among 1.0 x 10(6) leucocytes. The PCR assay revealed that normal bone marrow expresses low levels of EGP2 mRNA, although immunocytochemistry with the anti-EGP2 MAb MOC31 could not identify any positively stained cell. Analyses using this RT-PCR assay may prove to have applications to the assessment of circulating tumour cells in clinical samples.
authors.affiliation: Department of Clinical Immunology, University Hospital Groningen, The Netherlands.
authors.forename: W
authors.initials: W
authors.lastname: Helfrich
authors.forename: R
authors.initials: R
authors.lastname: ten Poele
authors.forename: G J
authors.initials: GJ
authors.lastname: Meersma
authors.forename: N H
authors.initials: NH
authors.lastname: Mulder
authors.forename: E G
authors.initials: EG
authors.lastname: de Vries
authors.forename: L
authors.initials: L
authors.lastname: de Leij
authors.forename: E F
authors.initials: EF
authors.lastname: Smit
mesh_terms: D000951:Antigens, Neoplasm / Q000032:analysis* / Q000235:genetics; D001853:Bone Marrow / Q000276:immunology; D001943:Breast Neoplasms / Q000097:blood*; D002277:Carcinoma / Q000097:blood*; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D006801:Humans; D009360:Neoplastic Cells, Circulating; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis; D012194:RNA-Directed DNA Polymerase; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
chemical_list: D000951:Antigens, Neoplasm; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D012333:RNA, Messenger; D012194:RNA-Directed DNA Polymerase