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Biological probe as well as preparation method and application thereof

A biological probe and probe technology, applied in the field of biochemistry, can solve problems such as insufficient specificity and sensitivity, false positive interference, low specificity, etc., and achieve the effects of improving specificity and sensitivity, improving recognition and combination, and simplifying detection components

Pending Publication Date: 2022-05-10
SHENZHEN INST OF ADVANCED TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Nucleic acid RNA detection generally uses RT-PCR technology to directly detect the presence of new coronavirus RNA in the sample, but it needs to manually extract the RNA virus in the sample, which is affected by multiple factors such as sample sampling method, sample transportation and storage, manual operation, etc., often accompanied by false negative interference , and a small number of RNA test results were positive and did not show infectivity, so there is also false positive interference; the detection of IgG / IgM antibodies to the new coronavirus is an indirect detection method, by detecting whether there are IgG / IgM antibodies to the new coronavirus in the blood To indirectly characterize whether the subject is infected with the new coronavirus, but it cannot indicate whether there are new coronavirus particles in the subject in real time, so it is impossible to judge whether the subject is infectious
[0004] In addition, in terms of wider virus detection, there are also reports on the use of sugar-gold nanoprobes for virus detection. However, in these reports, natural sialic acid ligands are generally used as detection molecules to modify the surface of gold nanoparticles to construct probes. However, the recognition and binding of natural sialic acid ligand molecules to virus surface proteins is a low-specificity and low-affinity recognition and binding effect. report

Method used

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  • Biological probe as well as preparation method and application thereof
  • Biological probe as well as preparation method and application thereof
  • Biological probe as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1: Synthesis of Custom Sialic Acid Ligand Molecule 1

[0067]

[0068] Natural sialic acid Neu5Ac (10g, 32.3mmol) was dissolved in ethanol (100mL), potassium carbonate (5.36g, 38.8mmol) was added, stirred for 15 minutes, benzyl bromide (6.08g, 35.6mmol) was added dropwise, stirred at 50°C After 2 hours, TLC detected that the reaction was complete, and the reaction solution was spin-dried, and the residue was spin-evaporated three times with toluene, and then drained to dryness to obtain the crude intermediate of formula 1-1, which was directly used in the next reaction;

[0069] The crude product of formula 1-1 obtained in the previous step was dissolved in chloroacetyl (20 mL), stirred at room temperature until the reaction was complete, and the solvent was evaporated to obtain the crude product of formula 1-2, which was directly used in the next reaction;

[0070] The crude product of formula 1-2 obtained in the previous step and HOPEG 4 OBn (13.79g, 48.5m...

Embodiment 2

[0076] Example 2: Synthesis of Custom Sialic Acid Ligand Molecule 2

[0077]

[0078] Concentrated hydrochloric acid (1 mL) was dropped into methanol (100 mL), natural sialic acid Neu5Ac (10 g, 32.3 mmol) was added in batches, the reaction was stirred until the solution was clear, the reaction solution was neutralized, the solvent was evaporated, and the residue was rotary evaporated three times by adding toluene , drained to obtain the crude intermediate of formula 2-1, which was directly used in the next step reaction;

[0079] The crude product of formula 2-1 obtained in the previous step was dissolved in chloroacetyl (40 mL), stirred at room temperature until the reaction was complete, and the solvent was evaporated to obtain the crude product of formula 2-2, which was directly used in the next reaction;

[0080] The crude product of formula 2-2 obtained in the previous step and HOPEG 4 OBn (13.8g, 48.5mmol) was mixed with dry dichloromethane (60mL), silver carbonate (...

Embodiment 3

[0092] Embodiment 3: Preparation of sialic acid-gold nanorod probe 3

[0093]

[0094] The custom-made sialic acid ligand molecule 1 obtained in Example 1 was dissolved in water to make an aqueous solution (10 mM), and 100 μL was added to the gold nanorod solution covered by CTABB, 75 nm in length, and with an aspect ratio of 3:1 (OD=15, 5 mL), after standing for 15 hours, centrifuged at 8000 rpm for 10 minutes, discarded the supernatant, and redispersed the centrifuge tube substrate aggregates in deionized water (5 mL) to obtain sialic acid-gold nanorod probe 3;

[0095] Wherein, the preparation process of gold nanorod solution comprises: adding chloroauric acid HAuCl 4 Add aqueous solution (0.5mM, 5mL) to CTAB aqueous solution (0.2M, 5mL), add fresh sodium borohydride NaBH under vigorous stirring 4 Ice-water solution (10mM, 600μL), continue to stir for 10 minutes to prepare gold nano-seed solution; HAuCl 4 Aqueous solution (5mM, 6mL), silver nitrate AgNO 3 Add aqueous ...

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Abstract

The invention discloses a biological probe as well as a preparation method and application thereof. The biological probe comprises a probe carrier and customized sialic acid ligand molecules connected with the probe carrier, the structural general formula of the customized sialic acid ligand molecule is as shown in formula I, wherein R1 is selected from hydroxyl, methoxyl or substituted methoxyl; r2 is selected from acetamido, substituted acetamido, benzamido, substituted benzoyl, alkoxycarbonylamido, triazolyl or substituted triazolyl, and R3 is selected from hydroxyl, methoxyl, substituted methoxyl, acetamido, substituted acetamido, sulfamido or phosphamido; n is selected from 0, 1, 2, 3, 4, 5, 6 and 7, and m is selected from 2, 3, 4, 5, 6, 7, 8 and 9. The biological probe can be used for directly detecting new coronavirus particles.

Description

technical field [0001] The invention belongs to the technical field of biochemistry, and relates to a biological probe and its preparation method and application. Background technique [0002] At present, the detection methods of the new coronavirus mainly include nucleic acid RNA detection and new coronavirus IgG / IgM antibody detection. The former extracts the nucleic acid substances in the sample to be tested and uses polymerase chain reaction (polymerase chain reaction, PCR) to amplify the RNA of the sample to be tested. To identify whether it is the RNA of the new coronavirus, real-time fluorescent quantitative PCR technology is generally used for detection; the latter uses the immune system to produce corresponding IgG and IgM antibodies after a person is infected with the new coronavirus, and detects the new coronavirus in human blood. Viral IgG and IgM are used to indirectly indicate whether the subject has a new coronavirus infection, and the colloidal gold method is...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/58G01N33/569G01N21/552C07H15/08C07H1/00B22F9/24B82Y30/00B82Y40/00B22F1/054
CPCG01N33/587G01N33/56983G01N21/554C07H15/08C07H1/00B22F9/24B82Y30/00B82Y40/00G01N2333/165
Inventor 李伟王怀雨
Owner SHENZHEN INST OF ADVANCED TECH
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