A quantitative method to measure telomerase activity by bioluminescence connected with telomeric repeat amplification protocol.
Telomerase is expected to be a new biomarker for cancer diagnosis. The telomeric repeat amplification protocol (TRAP) is a sensitive method to detect telomerase activity. However, TRAP and its modified protocols are not always suitable for measuring telomerase activity of a large number of clinical samples to diagnosis cancer because these methods generally require a time-consuming detection step such as gel electrophoresis. To improve the procedure for mass diagnosis, we applied bioluminescence to replace the detection step. Telomerase activity is measured by evaluating the amount of inorganic pyrophosphate generated in PCR amplification of telomerase elongation product, with use of the sensitive enzymatic luminometric inorganic pyrophosphate detection assay (ELIDA). TRAP connected with ELIDA (TRAP-ELIDA) can quantitatively detect telomerase activity within linearity from 2 to 1000 cell equivalents. The ELIDA signals accorded with results of TRAP-SYBR green staining, and the results of ELIDA were significantly correlated to those of TRAP connected with an enzyme-linked immunosorbent assay (TRAP-ELISA) (r(2) = 0.992, P < 0.001). TRAP-ELIDA is a simple and sensitive method to quantify telomerase activity without time-consuming gel electrophoresis. Because TRAP-ELIDA measures telomerase activity with a luminometer, it could be applied to a large number of clinical samples at the same time.
Xu
S
S
National Laboratory of Biomedical Photonics, Institute of Environmental Medicine, Tongji Medical College of Huazhong University of Science and Technology, 13 Hangkong Road, Wuhan 430030, People's Republic of China. shunqing@public.wh.hb.cn
He
M
M
Yu
H
H
Cai
X
X
Tan
X
X
Lu
B
B
Shu
B
B
D000255:Adenosine Triphosphate / Q000032:analysis; D002277:Carcinoma / Q000201:enzymology; D003838:Deoxyadenine Nucleotides / Q000737:chemistry; D005260:Female; D006801:Humans; D008156:Luciferases / Q000032:analysis; D008163:Luminescent Measurements; D016133:Polymerase Chain Reaction / Q000379:methods*; D012091:Repetitive Sequences, Nucleic Acid; D012680:Sensitivity and Specificity; D019098:Telomerase / Q000032:analysis*; D016615:Telomere / Q000235:genetics; D014407:Tumor Cells, Cultured
D003838:Deoxyadenine Nucleotides; D000255:Adenosine Triphosphate; D008156:Luciferases; D019098:Telomerase; C026600:2'-deoxyadenosine triphosphate