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A quantitative method to measure telomerase activity by bioluminescence connected with telomeric repeat amplification
1National 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
Analytical Biochemistry
|December 4, 2001
Summary
A new method, telomerase activity detection using the enzymatic luminometric inorganic pyrophosphate detection assay (ELIDA), simplifies cancer diagnosis. TRAP-ELIDA offers a fast, sensitive, and scalable approach for analyzing telomerase activity in clinical samples.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Telomerase is a key biomarker for cancer diagnosis.
- Traditional telomeric repeat amplification protocol (TRAP) methods are time-consuming for mass clinical sample analysis due to detection steps like gel electrophoresis.
Purpose of the Study:
- To develop a faster and more scalable method for measuring telomerase activity for cancer diagnosis.
- To replace the gel electrophoresis detection step in TRAP with a bioluminescence-based assay.
Main Methods:
- Applied the enzymatic luminometric inorganic pyrophosphate detection assay (ELIDA) to quantify inorganic pyrophosphate generated during PCR amplification of telomerase elongation products.
- Developed a combined method: TRAP-ELIDA.
- Validated TRAP-ELIDA against TRAP-SYBR green staining and TRAP-ELISA.
Main Results:
- TRAP-ELIDA quantitatively detects telomerase activity with linearity from 2 to 1000 cell equivalents.
- ELIDA signals correlated well with TRAP-SYBR green staining.
- ELIDA results showed a strong significant correlation with TRAP-ELISA (r(2) = 0.992, P < 0.001).
Conclusions:
- TRAP-ELIDA is a simple, sensitive, and rapid method for quantifying telomerase activity, eliminating the need for gel electrophoresis.
- The use of a luminometer allows TRAP-ELIDA to be applied to a large number of clinical samples simultaneously, facilitating mass diagnosis.