sybr-green-i and Urinary-Bladder-Neoplasms

To investigate whether hTERT mRNA expression could be used as a diagnostic and prognostic marker of bladder transitional cell carcinoma.. SYBR green I real-time fluorescence quantitative PCR method was used to quantify hTERT mRNA expression in exfoliated cells from patients with bladder transitional cell carcinomas.. A significant correlation was observed between the level of hTERT mRNA expression in exfoliated cells collected in urine with pathological grade and clinical stage.. Our results show that SYBR green real-time fluorescence quantitative evaluation of hTERT mRNA could be a useful marker of tumor progression for the early diagnosis and prognosis of bladder cancers.

Topics: Adult; Aged; Aged, 80 and over; Benzothiazoles; Biomarkers, Tumor; Carcinoma, Transitional Cell; Cells, Cultured; Diamines; Female; Gene Expression; Humans; Male; Middle Aged; Neoplasm Staging; Neoplastic Cells, Circulating; Organic Chemicals; Polymerase Chain Reaction; Prognosis; Quinolines; Reference Values; RNA, Messenger; Telomerase; Urinary Bladder Neoplasms

Tumor cells acquire the ability to enter blood vessels surrounding the primary tumor, endowing them with the capacity to disseminate and become established in distant sites, originating a metastasis. Determination of the intravasation ability of tumor cells is thus important, as it can be correlated with their potential malignancy. To analyze the intravasation phenotype of human tumor cells in vivo, we performed chick embryo chorioallantoic membrane (CAM) assays. Cells were inoculated on the CAM of 9-day-old chick embryos and the membrane at the opposite side of the egg was recovered after 48 h incubation. To measure intravasation ability, we calculated the amount of human DNA in each CAM sample by real-time PCR of Alu sequences and SYBR Green 1 fluorescence detection. This analysis showed a detection limit of 1 human cell per 10(5) total cells, and we were able to distinguish between tumor cells of distinct invasive capacity. This assay has several advantages over current methods to measure intravasation ability, including the elimination of post-PCR analysis, sensitivity and easy scale-up of sample numbers.

Topics: Adenocarcinoma; Allantois; Alu Elements; Animals; Benzothiazoles; Breast Neoplasms; Chick Embryo; Chorion; Computer Systems; Diamines; DNA, Neoplasm; Ethidium; Fibroblasts; Fluorescent Dyes; Humans; Neoplasm Invasiveness; Organic Chemicals; Phenotype; Polymerase Chain Reaction; Quinolines; Tumor Cells, Cultured; Urinary Bladder Neoplasms