PCR-ELISA telomerase assay
Telomere is a specific DNA-protein structure at the end of chromosomes in eukaryotes. Telomere DNA is a series of repeated G-rich DNA sequences. This sequence is highly conservative in biological evolution (human repeat sequence is TTAGGG). It has been confirmed that telomeres play an important role in protecting genomic DNA from degradation and preventing harmful binding of chromosomes (such as chromosome end fusion, rearrangement, chromosome shift, and chromosome deletion).
Since DNA polymerase cannot replicate the very end of linear DNA, the telomere ends of most normal somatic cells progressively shorten with each replication cycle. This phenomenon has been confirmed both in vivo and in vitro, and may be related to the limited reproductive capacity of normal somatic cells of higher eukaryotes.
This activity may play a role in conditions related to cell aging. In contrast to somatic cells, germ cells are immortal, and they need to preserve all genetic information for future organ formation. Therefore it must fight against telomere shortening. This work is done by adding a new telomere repeat sequence at the end of the genome chromosome.
Telomerase is a ribonucleoprotein. They use the complementary sequences of their own RNA components as templates to catalyze the addition of TTAGGG repeats to the ends of chromosomes. Telomerase activity is also expressed in most immortal cell lines and tumor cell lines. Telomerase reaction exceeds the limit of cell proliferation, which may be a prerequisite for the occurrence of malignant tumors.
Traditional telomerase research methods measure telomerase activity based on probe extension. Due to the large amount of sample materials required and the limited detection sensitivity, this method has been replaced by the Telomere Repeat Amplification Protocol (TRAP).
The standard TRAP assay is to amplify the telomerase reaction product by PCR, use a radioactive label, and after gel electrophoresis, display the results by autoradiography.
Here we introduce a new method for detecting telomerase using a non-radioactive label: active optical density enzyme-linked immunoassay. This method has the following characteristics:
Safe without using radioisotopes
One-step reaction, using ready-to-use mixtures for telomerase-mediated primer extension and PCR amplification can be performed in one test tube.
It is widely used and can be applied to different analysis methods after amplification.
Sensitive, comparable to the radioactive isotope TRAP measurement.
Reliable, accurate and repeatable.
Fast, get results in 6-8 hours.
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