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Preparation method for fluorescent western-blotting magnetic composite microballoon

A magnetic composite microsphere and fluorescent protein technology, which is applied in the fields of material science and engineering and biological separation engineering, can solve the problems of cumbersome and complicated operation process, and achieve the effects of fast mass transfer rate, high adsorption efficiency and uniform particle size.

Active Publication Date: 2015-01-14
YANTAI INST OF COASTAL ZONE RES CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Molecularly imprinted polymer microspheres without magnetic components used in the field of separation engineering are mostly used for static separation or dynamic separation, which need to be separated by centrifugation and other processes, and the operation process is cumbersome and complicated

Method used

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  • Preparation method for fluorescent western-blotting magnetic composite microballoon
  • Preparation method for fluorescent western-blotting magnetic composite microballoon
  • Preparation method for fluorescent western-blotting magnetic composite microballoon

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] a.Fe 3 o 4 Magnetic material preparation: weigh 3.00g of FeCl 3 ·6H 2 O was dissolved in 90mL of ethylene glycol, and 7.2g of sodium acetate was added, sonicated and mechanically stirred to dissolve. Then add 3.9g of polyethylene glycol and stir at 400r / min at 60°C until dissolved. After dissolution, all the yellow liquid was transferred to a closed hydrothermal reaction kettle with a polytetrafluoroethylene liner, and reacted in a dry oven at 200°C for 10 hours. After cooling to room temperature, the black substance was collected magnetically, washed with ultrapure water and absolute ethanol for 6 times, and the obtained product was dried for 24 hours, and the dried Fe 3 o 4 magnetic material.

[0043] b. Synthesis of superparamagnetic carrier particles by one-step miniemulsion polymerization: 1.0 g of the above-mentioned Fe 3 o 4 The magnetic material was mixed with 1 mL of oleic acid to obtain a black viscous gel. Methyl methacrylate (1.28 mL) and ethylene g...

Embodiment 2

[0052] Weigh 20mg of phycocyanin, dilute it in a 1L volumetric flask with redistilled water to obtain a 20mg / L phycocyanin solution, take 10mL of phycocyanin solution and add it to a 20mL screw-top vial, add 100mg of fluorescent Western blotting magnetic composite microspheres, oscillating for 10 minutes, and using a magnet to adsorb on one side of the vial for 2 minutes, the composite microspheres can be adsorbed to one side to obtain a clear solution (see Figure 2A ). It shows that the composite microsphere has a strong adsorption capacity for phycocyanin, and the separation process is fast and simple, see Figure 2B The magnetic strength of the microspheres is shown to be high.

Embodiment 3

[0054] Take 200 μL of 7 parts of 100 mg / L phycocyanin solution, place them in 10 mL plastic centrifuge tubes, and then add phosphate buffer solution (Na 2 HPO 4 and NaH 2 PO 4 0.2mol / L, volume ratio 1:1) 9.8mL, dilute to obtain 2mg / L phycocyanin solutions with pH values ​​of 5.6, 6.0, 6.4, 6.8, 7.2, 7.6 and 8.0, respectively, with a fluorescence spectrophotometer Determination of its corresponding fluorescence intensity, the maximum fluorescence intensity can be obtained when the pH value is 6.4 (see Figure 3A ), and as the concentration of phycocyanin increases, the fluorescence intensity increases gradually (see Figure 3B ).

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Abstract

The invention belongs to the field of material science and engineering and bioseparation engineering, and concretely relates to a preparation method for a fluorescent western-blotting magnetic composite microballoon. A miniemulsion polymerization method is employed to take phycocyanin as a molecular imprinting template and combine with a magnetically-sensitive fluorescent material, so that the phycocyanin molecular-imprinted-polymer magnetic fluorescent composite microballoon is obtained. By using the miniemulsion polymerization method, the molecular-imprinted-polymer microballoon integrates the magnetic and fluorescence double-responsive material, has specific recognition and autonomous adsorption on phycocyanin, is good in selectivity, high in adsorption efficiency, fast in speed, convenient and rapid for separation under the effect of an external magnetic field, and good in repeated usage performance, and is capable of realizing fluorescence real-time detection and fluorescence imaging at a relatively wide pH scope. Also the obtained magnetic core-shell blotting microballoon is regular in morphology and uniform in particle size. The preparation method has the advantages of low cost, simple and convenient operation, good reappearance and the like, and has wide application prospect.

Description

technical field [0001] The invention belongs to the fields of material science and engineering and biological separation engineering, and specifically relates to a preparation method of fluorescent protein imprinted magnetic composite microspheres. Background technique [0002] Molecularly imprinted technology refers to the process of preparing a polymer with specific selectivity for a specific target molecule, namely Molecularly Imprinted Polymer (MIP). "Technology. Due to the flexible structure and complex conformation of proteins, they are easily affected by environmental factors such as temperature. In addition, high cross-linking materials will prevent protein macromolecules from freely entering and leaving the polymer network. On the one hand, it makes it difficult to remove template molecules; The development of protein molecular imprinting materials has been hindered by a series of problems such as the difficulty of molecules entering the imprinting site and recombi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08F222/14C08F220/14C08F2/44C08F2/22C08F8/32C08F8/28C08J9/26C08K9/04C08K3/22B01J20/26B01J20/30
Inventor 陈令新张忠李金花
Owner YANTAI INST OF COASTAL ZONE RES CHINESE ACAD OF SCI
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