Method for detecting thrombin based on fluorescent dual-signal non-enzymatic amplification strategy of aptamer and application of method

Abstract

The invention belongs to the technical field of biomedicine, and particularly discloses a method for detecting thrombin based on a fluorescent dual-signal non-enzymatic amplification strategy of an aptamer and application of the method. The method comprises the following steps: reacting an auxiliary probe, a signal probe and a Linker chain to form a three-chain DNA reaction substrate, then mixingthe three-chain DNA reaction substrate, a fuel chain and the aptamer / catalyst chain for reaction to form a reaction solution, thus obtaining a thrombin fluorescent detection system, and after the thrombin is added, detecting the thrombin by monitoring the change of a fluorescent signal FAM and ROX. According to the method, the release of the catalyst chain is induced through the specific combination of the thrombin and the aptamer; the catalyst chain triggers and starts a DNA cyclic amplification reaction through nucleic acid hybridization and branch migration, so that the recycling and cyclic utilization and dual-signal amplification of the catalyst chain are realized; the whole reaction process of the method does not need any protease, and the whole reaction can be completed in one reaction system in one step; and the selectivity is good, the sensitivity is high, the linear range is 1fM-1nM, and the lowest detection lower limit is 0.45fM.

Description

technical field [0001] The invention relates to the field of biomedical technology, in particular to a method for detecting thrombin with a fluorescent double-signal enzyme-free amplification strategy based on nucleic acid aptamers and its application. Background technique [0002] The detection of proteins plays a vital role in the basic research and diagnosis of diseases. Thrombin, as a special serine protease, plays an important role in many life processes, such as blood coagulation, angiogenesis and wound healing process. To date, assays based on antigen-antibody recognition, including enzyme immunoassay and chemiluminescence, are the most commonly used methods for protein analysis. Due to the low abundance of proteins, the development of sensitive protein detection techniques and methods has become a bottleneck in the development of proteomics. [0003] Sensitivity and linear range are the main evaluation indicators of the detection method. The lower limit of detectio...

AI Technical Summary

Problems solved by technology

[0006] In view of the above-mentioned shortcomings of the prior art, the purpose of the present invention is to provide a method for detecting thrombin based on a nucleic acid aptamer-based fluorescence dual-signal enzyme-free amplification strategy and its application, which is used to solve the problem of thrombin in the prior art. The detection method has problems such as low sensitivity, narrow linear range, and slow detection speed

Images