Circulating nucleic acid detection kit based on microfluidic microbead array chip and application method thereof

Abstract

The invention belongs to the technical field of biomolecule detection, and discloses a circulating nucleic acid detection kit based on a microfluidic microsphere array chip and an application method thereof. The circulating nucleic acid detection kit based on the microfluidic microsphere array chip comprises a microfluidic detection chip for capturing probe functionalized microspheres, four LAMP primers and LAMP amplification reagents, a LAMP basic structure cyclization reagent and a cyclization auxiliary probe, a rolling ring amplification reagent, and a signal probe modified graphene quantumdot. The method is simple in operation and extremely high in sensitivity, and the early diagnosis of nasopharyngeal carcinoma can be achieved through detection of Epstein-Barr virus in blood. According to the circulating nucleic acid detection kit based on the microfluidic microsphere array chip and the application method thereof, the microscale and high-sensitivity detection characteristics of the micro-fluidic dynamic microarray chip, the multi-signal amplification (including loop-mediated isothermal amplification and rolling loop amplification) of the high-sensitivity target and the functional graphene quantum dot fluorescence signal amplification are integrated for the first time, and 10 mol / L Epstein-Barr virus DNA can be detected.

Description

technical field [0001] The invention belongs to the technical field of biomolecular detection, and in particular relates to a microfluidic microbead array chip-based circulating nucleic acid detection kit and an application method. Background technique [0002] At present, the existing technologies commonly used in the industry are as follows: [0003] Circulating tumor nucleic acid (ctDNA) has a low concentration in blood and there is a high level of interference from normal circulating DNA, so it is a very challenging task to establish a rapid and highly sensitive ctDNA detection method. In the past decade, many analytical techniques with great development potential have been proposed for the detection of ctDNA, especially from 2012 to the present, some new concepts and testing methods have been introduced into this field. For example: technology systems based on quantitative PCR (such as Amplification RefractoryMutation System (ARMS), SNPase-ARMS qPCR, mutant-enriched PC...

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Problems solved by technology

[0006] (1) There are certain defects in these detection methods in the prior art, such as: the need for expensive instruments and equipment, high false positives, high detection cost, long analysis cycle, cumbersome experimental process, etc., are only suitable for high-risk areas (such as China Hong Kong, Guangdong and other regions) carry out screening among people, and it is simply difficult for ordinary people in low-risk areas to afford a series of high costs brought about by testing;

[0007] (2) Due to the shortcomings of low sensitivity and large sample volume, the existing conventional technology is difficult to develop into a clinical detection technology, especially it is difficult to detect target DNA with extremely low concentration in blood samples

[0008] (3) It is difficult to solve the difficult problem that high-throughput and high-sensitivity properties are difficult to coexist under the condition of micro-sample in the field of micro-medicine analysis technology or the field of minimally invasive and non-invasive diagnosis

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