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Immune lipid-polymer hybridized nanoparticle biological chip and preparation method and application thereof in disease detection

A biochip, polymer technology, applied in measurement devices, instruments, scientific instruments, etc., can solve the problems of low sensitivity, no fluorescence signal amplification, no cell membrane fusion, etc., and achieve high selectivity and sensitivity. Effect

Inactive Publication Date: 2017-05-31
胡家铭 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Immunoliposomes can specifically capture exosomes in blood and fuse with them into larger nanoscale complexes, but the "onion" structure of liposomes does not have the function of fluorescent signal amplification, which is very important for detection Exosomes containing one or a small number of target RNAs with low sensitivity in early cancer
Polymer nanoparticles have good mechanical stability, have a "core-shell" structure, and have a high encapsulation efficiency, but they do not have the characteristics of fusion with cell membranes
At present, there is no technical report on lipid-polymer hybrid nanoparticle biochip, and there is no technical report on lipid-polymer hybrid nanoparticle biochip for early detection of cancer

Method used

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  • Immune lipid-polymer hybridized nanoparticle biological chip and preparation method and application thereof in disease detection
  • Immune lipid-polymer hybridized nanoparticle biological chip and preparation method and application thereof in disease detection
  • Immune lipid-polymer hybridized nanoparticle biological chip and preparation method and application thereof in disease detection

Examples

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Effect test

Embodiment 1

[0074]Preparation and physical and chemical properties characterization of lipid-polymer hybrid nanoparticle biochip.

[0075] Example 1 describes a simple method to prepare a lipid-polymer hybrid nanoparticle biochip ( Figure 1a ,b). A gold-coated glass substrate was immersed in an ethanol solution of βME and SH-PEG-Biotin to form a self-assembled monolayer. The PDMS template was immobilized on the self-assembled monolayer by microcontact paste technology to make a chip. The hairpin DNA catalytic loop (H1, H2, and Reporter) is encapsulated in lipid-polymer hybrid nanoparticles, which are anchored to the chip surface by biotin-avidin-specific affinity. Cationic lipid-polymer hybrid nanoparticles capture anionic exosomes through electrostatic interaction and fuse with them to form nanoscale complexes, so that the hairpin DNA catalytic circuit (CHDC) in nanoparticles and the exosomes in exosomes The target RNA hybridizes and produces an amplified fluorescent signal. The targ...

Embodiment 2

[0078] Lipid-polymer hybrid nanoparticle biochip for artificial exosome detection.

[0079] In this example, artificial exosomes ( Figure 2a ). Based on the fact that natural exosomes contain a small amount of target RNA and other RNAs, our artificial exosomes contain 1% GPC DNA and 99% miR54-DNA. Figure 2b -i shows the comparison of the sensitivity results of the detection of artificial exosomes by liposome biochips and lipid-polymer hybrid nanoparticle biochips respectively encapsulating molecular beacons and hairpin DNA catalytic circuits (CHDC). The results showed that the sensitivity of the lipid-polymer hybrid nanoparticle biochip wrapped with the hairpin DNA catalytic circuit (CHDC) was 600 times higher than that of the liposome biochip wrapped with molecular beacons, showing higher sensitivity.

Embodiment 3

[0081] Lipid-polymer hybrid nanoparticle biochip for detection of GPC1mRNA and KRAS in pancreatic cancer cells G12D mRNA.

[0082] Example 3 includes the application of lipid-polymer hybrid nanoparticle biochips containing molecular beacons or hairpin DNA catalytic circuits (CHDC) to distinguish pancreatic cancer cells (AsPC-1) from pancreatic normal cells (HPDE6-C7), And compared with liposome biochip. GPC1mRNA and KRAS G12D mRNA is an effective biomarker for pancreatic cancer cells and pancreatic cancer mutant cells, respectively. Figure 3a -d demonstrates that encapsulation of the hairpin DNA catalytic circuit (CHDC) targeting GPC1 mRNA can effectively differentiate pancreatic cancer cells from normal pancreatic cells. Figure 3e -f show package for KRAS G12D The hairpin DNA catalytic circuit (CHDC) of mRNA can effectively distinguish pancreatic cancer mutant cells from normal pancreatic cells.

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Abstract

The invention discloses an immune lipid-polymer hybridized nanoparticle biological chip and a preparation method and an application thereof in disease detection. The immune lipid-polymer hybridized nanoparticles are distributed on a chip in a form of a microarray. By taking a mixture of lipid and a polymer as a shell material, molecular probes with fluorescent labels are embedded thereinto. The immune lipid-polymer hybridized nanoparticles load various ligands on the surfaces. The biological chip can specifically capture exosomes in blood and is mixed with the same. Under the circumference of not damaging the exosomes, messenger RNAs and / or small molecular RNAs in the exosomes are detected and a fluorescence signal is generated, and meanwhile, the fluorescence signal is amplified. The chip disclosed by the invention has relatively high selectivity and sensitivity in detecting the exosomes containing one or a small amount of target RNAs in the early stage of cancers. Meanwhile, the biological chip has the advantages of being fast, simple, sensitive, high in selectivity, low in cost and the like, and can be used for molecular detection in the field of various diseases, biological and food safety and the like.

Description

technical field [0001] The invention relates to a biological chip and a preparation method thereof, more particularly to an immune lipid-polymer hybrid nanoparticle biochip, a preparation method thereof and an application thereof in disease detection. Background technique [0002] Exosomes are tiny membrane vesicles secreted by cells, with a size of 50-150nm, having a lipid bilayer membrane structure, and can stably exist in blood. Studies have shown that the concentration of exosomes in the blood of cancer patients is higher than that of normal people. In recent years, it has been found that exosomes contain a variety of functional molecules, such as: messenger RNA (mRNA), small molecule RNA (microRNA), DNA fragments, protein and peptides, etc. Given the importance of mRNA and microRNA in regulating gene expression and the dysfunction of their mutants in human diseases, they have become effective biomarkers for cancer detection. Quantitative determination of target mRNA a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574
CPCG01N33/57484
Inventor 胡家铭盛燕
Owner 胡家铭
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