Label-free ratio fluorescence type antigen detection method

A label-free, fluorescent technology, applied in the direction of measuring devices, biological testing, material inspection products, etc., can solve the problems of time-consuming steps, low sensitivity, high cost, etc., to achieve cost saving, high sensitivity, and avoid the effect of labeling secondary antibodies

Pending Publication Date: 2022-06-07
SOUTHWEST UNIVERSITY
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Problems solved by technology

[0007] The object of the present invention is to provide a label-free ratio fluorescence detection antigen method, which is used to provide a label-free, ratio method, an

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  • Label-free ratio fluorescence type antigen detection method

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Embodiment Construction

[0030] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, but not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0031] Aptamers are synthetic oligonucleotide or peptide sequences that fold into secondary and tertiary structures that bind to their corresponding targets with high affinity and specificity. Aptamers are more promising alternatives to antibodies due to their unique properties. First, the aptamers obtained by in vitro screening are easy to synthesize, which overcomes the limitation of using cell lines or animals to prepare antibodies...

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Abstract

The invention discloses a label-free ratio fluorescence type antigen detection method. The method comprises the following steps: designing a single-stranded DNA which comprises a recognition DNA sequence for CRISPR and an aptamer of a target antigen, and constructing a component A by spacing the recognition DNA sequence and the aptamer through a plurality of continuous T basic groups; by taking a target antigen as a target, modifying an antibody of the target antigen on a 96-well plate to construct a component B; connecting the surface-carboxylated silicon dioxide microspheres with a DNA chain of which the tail end is modified with amino groups, and complementarily pairing auxiliary DNA with DNA with amino groups to form double chains, so as to construct a component C; adding the component A, the component B, the component C, copper ions, ascorbic acid, DAPI dye and exonuclease III into an object to be detected; and detecting red fluorescence and blue fluorescence of an object to be detected, and detecting the target antigen by a ratio fluorescence method. According to the ratio fluorescence method, the intensities of two kinds of fluorescence are compared to serve as signal parameters, the accuracy is higher, the sensitivity is higher, and the selectivity is better; and moreover, the sensitivity is improved, the defect that a traditional method needs to label an antibody is overcome, and the detection cost is saved.

Description

technical field [0001] The invention relates to the technical field of antigen detection, in particular to a label-free ratio fluorescence detection method for antigen. Background technique [0002] As the gold standard for immunoassays, enzyme-linked immunosorbent assay (ELISA) is an extremely powerful technology for complex substances, which has been widely used in various fields, and a large number of commercial kits are also available. [0003] Traditional ELISA mainly relies on a specific antigen-antibody immune reaction, generally combined with a natural enzyme (horseradish peroxidase, HRP) catalyzed substrate as a colorimetric signal. There is no doubt that ELISA is excellent for detecting protein targets in biomedical and other fields. [0004] Nevertheless, traditional ELISA also has some disadvantages, such as low sensitivity, high cost, time-consuming and cumbersome steps. Typical ELISA assay concentrations are above the picomolar range. However, in the early s...

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Application Information

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IPC IPC(8): G01N33/68G01N33/552G01N33/58
CPCG01N33/68G01N33/552G01N33/582
Inventor 凌玉罗红群李念兵
Owner SOUTHWEST UNIVERSITY
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