Gold magnetic nanoprobe based on ordered arrangement of aptamers and application of gold magnetic nanoprobe in okadaic acid detection

An orderly arrangement technology of Datian dalsangic acid, applied in the field of analytical chemistry, can solve the problems of different conclusions, lodging of aptamers, inability to arrange orderly, etc., so as to improve the reactivity, improve the utilization efficiency, and reduce the synthesis cost. Effect

Pending Publication Date: 2022-06-24
FUZHOU UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the traditional gold-mercapto self-assembly method cannot arrange the density and configuration of DNA on the surface of gold nanoparticles in an orderly manner, and the aptamer lodging phenomenon often occurs, which is also the reason why researchers have studied DNA in nano-gold. One of the important reasons for the different conclusions obtained when gold surface reactivity

Method used

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  • Gold magnetic nanoprobe based on ordered arrangement of aptamers and application of gold magnetic nanoprobe in okadaic acid detection
  • Gold magnetic nanoprobe based on ordered arrangement of aptamers and application of gold magnetic nanoprobe in okadaic acid detection
  • Gold magnetic nanoprobe based on ordered arrangement of aptamers and application of gold magnetic nanoprobe in okadaic acid detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] A preparation method of gold magnetic nanoprobes (Apt@AuNPs@Fe3O4) with ordered arrangement of aptamers, the specific steps are as follows:

[0040] (1) Preparation of PEI-iron tetroxide nanoparticles:

[0041] Weigh 0.68g of ferric chloride hexahydrate (FeCl 3 ·6H 2 O) dissolved in 12 mL of ethylene glycol ((CH 2 OH) 2 ) to prepare solution A; weigh 0.5 g polyethyleneimine (PEI) and dissolve it in 4 mL of ethylene glycol to prepare solution B; under magnetic stirring (60°C), slowly add solution B to solution A , add 1.8 g of sodium acetate (CH 3 COONa), continue to stir for 10mins to obtain solution C; transfer the obtained solution C to the hydrothermal reaction kettle for reaction, the hydrothermal reaction kettle is placed in an oven and heated at a temperature of 220°C for 2 hours; the reaction kettle is naturally cooled to room temperature Afterwards, the solid phase was ultrasonically cleaned three times with deionized water and absolute ethanol in sequence,...

Embodiment 2

[0049] A method for preparing gold magnetic nanoprobes with disordered arrangement of aptamers, the specific steps are as follows:

[0050] (1) Preparation of PEI-iron tetroxide nanoparticles:

[0051] Weigh 0.68g of ferric chloride hexahydrate (FeCl 3 ·6H 2 O) dissolved in 12 mL of ethylene glycol ((CH 2 OH) 2 ) to prepare solution A; weigh 0.5 g polyethyleneimine (PEI) and dissolve it in 4 mL of ethylene glycol to prepare solution B; under magnetic stirring (60°C), slowly add solution B to solution A , add 1.8 g of sodium acetate (CH 3 COONa), continue to stir for 10mins to obtain solution C; transfer the obtained solution C to the hydrothermal reaction kettle for reaction, the hydrothermal reaction kettle is placed in an oven and heated at a temperature of 220°C for 2 hours; the reaction kettle is naturally cooled to room temperature Afterwards, the solid phase was ultrasonically cleaned three times with deionized water and absolute ethanol in sequence, and after magne...

Embodiment 3

[0059] Quantitative detection of OA by magnetic separation sensing system of ordered / disordered aptamers: 0.48 mg of gold magnetic nanoprobes prepared in Example 1 and Example 2 were selected respectively, and 40 μL of different concentrations were added to them. The OA solution was subjected to a competitive reaction at room temperature for 60 mins, and the supernatant was collected by magnetic separation. Each sample was tested three times and the average value was taken. The fluorescence measurement conditions were: capillary electrochromatography-laser-induced fluorescence (LIF) combined Detection platform, excitation wavelength is 497nm, emission wavelength is 512nm, chromatographic column size is 100μm × 500mm quartz capillary, mobile phase: PBS buffer with pH = 7.5, flow rate: 0.2mL / min, detection sample volume: 0.06-0.1μL , the PBS buffer is composed of 10 mmol / L NaH 2 PO 4 , 10mmol / L Na 2 HPO 4 , 500 mmol / L NaCl.

[0060] figure 2 It is a comparison of the effec...

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Abstract

The invention discloses a gold magnetic nanoprobe based on ordered arrangement of aptamers and application of the gold magnetic nanoprobe in okadaic acid detection (OA). The gold magnetic nanoprobe is formed by combining nanogold spherical nucleic acid and nano magnetic beads, wherein nucleic acid sequences are orderly arranged on the surface of the nanogold spherical nucleic acid; the nucleic acid sequence is formed by an aptamer with an adenine base polymeric chain at the 5'end and a DNA complementary chain with a fluorescent label at the 5 'end. The surface of the gold magnetic nanoprobe is modified with high density and is loaded with orderly arranged aptamers, so that the gold magnetic nanoprobe is high in specific recognition efficiency and simple and convenient to separate a target object, and can react with okadaic acid to release a fluorescently-labeled DNA complementary chain, and laser-induced fluorescence detection is applied, so that high-sensitivity analysis of trace OA is realized.

Description

technical field [0001] The invention relates to the technical field of analytical chemistry, in particular to a gold magnetic nano-probe with an orderly arrangement of nucleic acid aptamers and its application in the detection of Datian haisalic acid. Background technique [0002] Okadaic acid (OA) is a heat-stable fat-soluble polyether marine toxin, the most important one of diarrheal shellfish toxins, its molecular formula is C 44 H 88 O 13 , the molecular weight is 805.02. OA is originally produced by algae such as Pinnella and Prototheca, and is passed through the food chain to shellfish such as oysters, clams, and mussels, where it accumulates in their digestive glands. OA has no toxic hazard to these shellfish, but when people eat these poisonous shellfish, it will cause gastrointestinal disorders, such as diarrhea, abdominal pain, nausea, vomiting and other symptoms, which is called diarrheal shellfish poisoning. Therefore, the establishment of a new rapid and sen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/115G01N33/53G01N27/447G01N21/64
CPCC12N15/115G01N33/5308G01N27/447G01N21/6402C12N2310/16
Inventor 林旭聪黄洋童诗谦谢增鸿
Owner FUZHOU UNIV
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