CN-121978325-A - Fluorescence RNA functionalized hydrogel kit, preparation method and method for detecting colorectal cancer miRNA-126

Abstract

The invention relates to a fluorescent RNA functionalized hydrogel kit, a preparation method and a method for detecting colorectal cancer miRNA-126, which solve the technical problem of how to detect miRNA-126 in blood or tissue samples, are suitable for portable detection, the fluorescent RNA functionalized hydrogel is obtained by crosslinking methacrylic acid hyaluronic acid, an acryl modified fluorescent RNA aptamer and N, N' -methylenebisacrylamide, and covalent crosslinking between the fluorescent RNA aptamer and the hydrogel can prevent the RNA aptamer from being released into an external environment, so that the stability and the anti-interference capability of the hydrogel kit are improved.

Inventors

• Liu Qianrui
• ZOU LINGLONG

Assignees

• 温州康瑞佰欧生物技术有限公司

Dates

Publication Date

20260505

Application Date

20260106

Claims (10)

1. 1. A fluorescent RNA functionalized hydrogel kit comprising a fluorescent RNA functionalized hydrogel and a non-fluorescent dye.
2. 2. The fluorescent RNA functionalized hydrogel kit of claim 1, wherein the fluorescent RNA functionalized hydrogel comprises a dissolution solution, a methacryloylated hyaluronic acid, a bisacrylamide, a photoinitiator, and fluorescent RNA, the methacryloylated hyaluronic acid, bisacrylamide, photoinitiator, and fluorescent RNA cross-linked.
3. 3. The fluorescent RNA functionalized hydrogel kit of claim 1, wherein the non-fluorescent dye is HBC 530.
4. 4. The fluorescent RNA functionalized hydrogel kit of claim 1, wherein the fluorescent RNA is an acryl modified fluorescent RNA.
5. 5. A method of detecting colorectal cancer miRNA-126 using the fluorescent RNA-functionalized hydrogel kit of claim 1, comprising the steps of: step S1, preparation of miRNA-126 detection solution: preparing a sample to be detected into a solution in double distilled water without RNase to form a sample to be detected; S2, incubating the fluorescent RNA functionalized hydrogel with miRNA-126 detection solution; And S3, collecting fluorescent signals by a fluorescent microscope, and identifying miRNA-126.
6. 6. A method for detecting colorectal cancer miRNA-126 using the fluorescent RNA-functionalized hydrogel kit of claim 2, comprising the steps of: step S1, preparation of miRNA-126 detection solution: preparing a sample to be detected into a solution in double distilled water without RNase to form a sample to be detected; S2, incubating the fluorescent RNA functionalized hydrogel with miRNA-126 detection solution; And S3, collecting fluorescent signals by a fluorescent microscope, and identifying miRNA-126.
7. 7. The method of claim 6, wherein the concentration of miRNA-126 in the sample is calculated from the fluorescent signal.
8. 8. A method of preparing a fluorescent RNA functionalized hydrogel kit of claim 2, wherein the fluorescent RNA functionalized hydrogel is prepared by: Adding the photoinitiator and the bisacrylamide into a dissolving solution to dissolve the photoinitiator, then adding the methacryloyl hyaluronic acid and the acryl modified fluorescent RNA into the solution to obtain a mixed solution, dripping the mixed solution into an immunofluorescence glass-carrying plate hole, and exposing to achieve crosslinking.
9. 9. A fluorescent RNA functionalized hydrogel, comprising a dissolution solution, a methacryloylated hyaluronic acid, a bisacrylamide, a photoinitiator, and a fluorescent RNA, wherein the methacryloylated hyaluronic acid, the bisacrylamide, the photoinitiator, and the fluorescent RNA are crosslinked.
10. 10. The fluorescent RNA functionalized hydrogel kit of claim 2, wherein the mass ratio of bisacrylamide to methacryloylated hyaluronic acid is 1:4.

Description

Fluorescence RNA functionalized hydrogel kit, preparation method and method for detecting colorectal cancer miRNA-126 Technical Field The invention relates to the technical field of biological detection, in particular to a fluorescence RNA functionalized hydrogel kit, a preparation method and a method for detecting colorectal cancer miRNA-126. Background Colorectal cancer is the third most common cancer worldwide, with high morbidity and mortality, and is always a public health problem. Colonoscopy is recognized clinically as a "gold standard" for diagnosing colorectal cancer. However, colonoscopy is invasive, complex and time consuming, with the risk of bleeding and perforation, and long-term monitoring by colonoscopy faces the problem of poor patient compliance. Therefore, there is an increasing need for non-invasive diagnostic methods in colorectal cancer screening. Due to the advantages of non-invasive acquisition and low cost, cancer-related biomarkers are well suited for a wide range of applications in clinical practice. Micrornas (mirnas) play an important role in gene expression and immune-mediated diseases. Related studies have shown that miRNA-126 is capable of inhibiting the growth, invasion, metastasis and epithelial-to-mesenchymal transition of colon cancer cells, suggesting that miRNA-126 is a potential cancer-associated biomarker. It can be seen that by quantifying the expression level of miRNA-126 in blood or tissue samples, key evidence can be obtained to support early detection or monitoring of colorectal cancer. Therefore, how to detect miRNA-12 in blood or tissue samples is a technical problem to be solved by those skilled in the art. Disclosure of Invention The application provides a fluorescence RNA functionalized hydrogel kit, a preparation method and a method for detecting colorectal cancer miRNA-126, and aims to solve the technical problem of how to detect miRNA-126 in blood or tissue samples. Fluorescent RNA aptamers, also known as fluorescent "lighting" aptamers, are capable of activating the fluorescence of non-fluorescent dyes by folding into a specific structure. Fluorescent RNA aptamer is a powerful tool for visual detection of RNA, and has the advantages of high quantum yield, rapid fluorescence activation, small size, good photobleaching resistance and the like. More importantly, the fluorescent RNA sensor has lower background and higher signal to noise ratio, so that the fluorescent RNA sensor has wide and promising application in the field of visual fluorescent sensing. Hydrophilic hydrogels have advantages in terms of loading capacity and mechanical stability due to their porous polymer network structure, and can uniformly fix fluorescent RNAs in pores. In a first aspect of the present disclosure, a fluorescent RNA functionalized hydrogel kit is provided, comprising a fluorescent RNA functionalized hydrogel and a non-fluorescent dye. Preferably, the fluorescent RNA functionalized hydrogel comprises a solution, methacryloylated hyaluronic acid, bisacrylamide, a photoinitiator, and fluorescent RNA, the methacryloylated hyaluronic acid, bisacrylamide, a photoinitiator, and fluorescent RNA cross-links. Preferably, the non-fluorescent dye is HBC 530. Preferably, the fluorescent RNA is an acryl-modified fluorescent RNA. In a second aspect of the present disclosure, there is provided a method of detecting colorectal cancer miRNA-126 using a fluorescent RNA-functionalized hydrogel kit, comprising the steps of: step S1, preparation of miRNA-126 detection solution: preparing a sample to be detected into a solution in double distilled water without RNase to form a sample to be detected; S2, incubating the fluorescent RNA functionalized hydrogel with miRNA-126 detection solution; And S3, collecting fluorescent signals by a fluorescent microscope, and identifying miRNA-126. Preferably, the concentration of miRNA-126 in the sample is calculated from the fluorescence signal. In a third aspect of the present disclosure, there is provided a method of preparing a fluorescent RNA functionalized hydrogel kit by: Adding the photoinitiator and the bisacrylamide into a dissolving solution to dissolve the photoinitiator, then adding the methacryloyl hyaluronic acid and the acryl modified fluorescent RNA into the solution to obtain a mixed solution, dripping the mixed solution into an immunofluorescence glass-carrying plate hole, and exposing to achieve crosslinking. In a fourth aspect of the present disclosure, there is provided a fluorescent RNA functionalized hydrogel comprising a dissolution solution, methacryloylated hyaluronic acid, bisacrylamide, a photoinitiator, and fluorescent RNA, the methacryloylated hyaluronic acid, bisacrylamide, photoinitiator, and fluorescent RNA crosslinked. The kit has the beneficial effects that the kit can effectively, qualitatively and quantitatively detect miRNA-126, the linear range is 0.1-10000 nM, and the detection limit is 23.8 pM. And visual detection i