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A colorimetric detection method for telomerase activity

A technology of telomerase and detection method, applied in the field of tumor detection, can solve the problems of low practicability and expensive instruments, etc.

Active Publication Date: 2020-04-24
SHAANXI NORMAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The technical problem to be solved by the present invention is to overcome the defects of expensive instruments required for telomerase activity detection in the prior art and poor practicability, and to provide a method based on horseradish peroxidase (HRP) to catalyze 3,3', Oxidation of 5,5'-Tetramethylbiphenyl for a Simple and Sensitive Colorimetric Detection of Telomerase Activity

Method used

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  • A colorimetric detection method for telomerase activity
  • A colorimetric detection method for telomerase activity
  • A colorimetric detection method for telomerase activity

Examples

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Embodiment 1

[0022] Taking the detection of telomerase in HeLa cells (HeLa) as an example, the specific detection method is as follows:

[0023] 1. Add 3.2 μL of aminated magnetic beads (NH 2 -MB, produced by Shaanxi North American Gene Co., Ltd.) was washed 3 times with 0.1mol / L PBS buffer solution, and then the cleaned aminated magnetic beads were added to 360 μL 0.1mol / L PBS buffer solution, and 40 μL concentration of 2 μmol / L aldehyde-formylated telomerase primer (provided by Sangon Bioengineering (Shanghai) Co., Ltd., sequence: 5'-CHO-TTTTTTTTTTTTTTTTTTTTTAATCCGTCGAGCAGAGTT-3') was reacted at 30°C for 2 hours by vortexing. After the reaction, wash twice with an equal volume of 0.1mol / L PBS-T buffer solution and 0.1mol / L PBS buffer solution, then disperse the washed magnetic beads in 400 μL 0.1mol / L PBS buffer solution, and send to Add 25.2 μL of n-valeraldehyde aqueous solution with a concentration of 7 mmol / L, and react with vortex shaking at 25° C. for 10 hours. After the reaction...

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Abstract

The invention discloses a telomerase-activity colorimetric detection method. Naked-eye colorimetric detection of telomerase activity is realized with the method combined with magnetic separation and by the aid of catalytic action of horse radish peroxidase on 3, 3', 5, 5'-tetramethyl benzidine; when active telomerase exists, telomerase substrates modified on the surfaces of magnetic beads can be extended, the extended substrates can be hybridized with cDNA (complementary deoxyribonucleic acid) of multiple 5'end modified biotins, the cDNA hybridized to the surfaces of the magnetic beads can be specifically integrated with streptavidin-horse radish peroxidase through specific action of the avidin and the biotins, color of a solution is enabled to change through oxidation of the 3, 3', 5, 5'-tetramethyl benzidine catalyzed by the horse radish peroxidase, and simple and flexible naked-eye color detection of the telomerase activity can be realized. Expression level of the telomerase activity of different types of cancer cells can be detected, the cancel cells are distinguished from normal cells, and great significance in therapeutic monitoring of cancer detection by taking the telomerase as markers and the telomerase as targets is achieved.

Description

technical field [0001] The invention belongs to the technical field of tumor detection, and in particular relates to a simple and sensitive naked-eye colorimetric detection method for telomerase activity in cancer cells based on the color change of the solution. Background technique [0002] Telomere is a special structure located at the end of chromosomes in eukaryotic cells. It is composed of telomere DNA and telomere-related proteins. The telomere DNA is composed of non-coding repeat sequences. Different species have different repeat sequences, such as human telomere The DNA is composed of 5'-TTAGGG-3' repeat sequence, and the length ranges from 4 to 15kb. The main function of telomeres is to protect the ends of chromosomes, prevent chromosome fusion, recombination and degradation. In normal cells, as the cells proliferate, the telomeres will gradually shorten. When the telomeres shorten to a certain extent, the chromosomes will become unstable, and the cells will age an...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6816C12Q1/6886C12Q1/48
CPCC12Q1/485C12Q1/6816C12Q1/6886C12Q2326/12C12Q2600/158C12Q2563/125C12Q2521/113C12Q2563/143C12Q2563/149
Inventor 金燕王要才
Owner SHAANXI NORMAL UNIV
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