title: A rapid biosensor chip assay for measuring of telomerase activity using surface plasmon resonance.
abstract: Considerable interest has been focused on telomerase because of its potential use in assays for cancer diagnosis, and for anti-telomerase drugs as a strategy for cancer chemotherapy. A number of assays based on the polymerase chain reaction (PCR) have been developed for evaluation of telomerase activity. To overcome the disadvantages of the conventional telomerase assay [telomeric repeat amplification protocol (TRAP)] related to PCR artifacts and troublesome post-PCR procedures, we have developed a telomeric repeat elongation (TRE) assay which directly measures telomerase activity as the telomeric elongation rate by biosensor technology using surface plasmon resonance (SPR). 5'-Biotinylated oligomers containing telomeric repeats were immobilized on streptavidin-pretreated dextran sensor surfaces in situ using the BIACORE apparatus. Subsequently, the oligomers associated with the telomerase extracts were elongated in the BIACORE apparatus. The rate of TRE was calculated by measuring the SPR signals. We examined elongation rates by the TRE assay in 18 cancer and three normal human fibroblast cell lines, and 12 human primary carcinomas and matching normal tissues. The elongation rates increased in a concentration- and time-dependent manner. Those of cancer cells were two to 10 times higher than fibroblast cell lines and normal tissues. Telomerase activities and its inhibitory effects of anti-telomerase agents as measured by both the TRE and TRAP assays showed a good correlation. Our assay allows precise quantitative comparison of a wide range of human cells from somatic cells to carcinoma cells. TRE assay is suitable for practical use in the assessment of telomerase activity in preclinical and clinical trials of telomerase-based therapies, because of its reproducibility, rapidity and simplicity.
authors.affiliation: Department of Pathology, Iwate Medical University School of Medicine, Uchimaru 19-1, Morioka 202-8505, Japan. chihaya@iwate-med.ac.jp
authors.forename: Chihaya
authors.initials: C
authors.lastname: Maesawa
authors.forename: Toru
authors.initials: T
authors.lastname: Inaba
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Sato
authors.forename: Sin
authors.initials: S
authors.lastname: Iijima
authors.forename: Kaoru
authors.initials: K
authors.lastname: Ishida
authors.forename: Masanori
authors.initials: M
authors.lastname: Terashima
authors.forename: Ryo
authors.initials: R
authors.lastname: Sato
authors.forename: Michihiro
authors.initials: M
authors.lastname: Suzuki
authors.forename: Akiko
authors.initials: A
authors.lastname: Yashima
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ogasawara
authors.forename: Hiroki
authors.initials: H
authors.lastname: Oikawa
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Sato
authors.forename: Kazuyoshi
authors.initials: K
authors.lastname: Saito
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Masuda
mesh_terms: D002460:Cell Line; D004965:Estrogen Antagonists / Q000494:pharmacology; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics*; D015203:Reproducibility of Results; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology; D020349:Surface Plasmon Resonance / Q000379:methods*; D013629:Tamoxifen / Q000494:pharmacology; D019098:Telomerase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016615:Telomere / Q000235:genetics* / Q000378:metabolism; D014407:Tumor Cells, Cultured; D015215:Zidovudine / Q000494:pharmacology
chemical_list: D004965:Estrogen Antagonists; D018894:Reverse Transcriptase Inhibitors; D013629:Tamoxifen; D015215:Zidovudine; D019098:Telomerase