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Nano magnetic bead for nucleic acid purification

A nano-magnetic bead and magnetic bead technology, applied in the field of material science, can solve the problems of incomplete modification of surface groups, low degree of automation, and instability of magnetic beads, so as to reduce workload, realize automation, broad application prospects and social The effect of economic benefits

Inactive Publication Date: 2016-05-11
BEIJING NAGENE DIAGNOSTIC REAGENT CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The technical problem to be solved by the present invention is to provide a dispersing magnetic bead for the existing disadvantages of instability, incomplete modification of surface groups, need for organic solvent dispersion, irregular particles, small adsorption rate, and low degree of automation. Nano magnetic beads for nucleic acid purification with good stability, low cost, good particle uniformity, and strong controllability of magnetic bead size

Method used

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  • Nano magnetic bead for nucleic acid purification

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Experimental program
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Embodiment 1

[0037] Embodiment 1: Preparation of nano magnetic beads for nucleic acid purification

[0038] Step A) preparation of nano-microspheres:

[0039] Under the protection of argon, 1 ml of 0.5 M sodium hydroxide solution was added dropwise at intervals of 10 minutes to the aqueous solution containing 15 g of ferrous sulfate heptahydrate and 10 g of ferric chloride hexahydrate. When the reaction solution turns into a colloidal emulsion, add 1ml of 0.25M sodium hydroxide dropwise every 20 minutes, shake the shaker continuously at a speed of 100 rpm, and continue argon protection after the magnetic field appears. Under reaction 4~10 hours. Then, according to the volume of magnetic beads, sodium chloride was added to make the final concentration 1M. After standing at room temperature for 18-36 hours, rinse with deionized water until the pH of the solution is close to 7. Use 0.25M sodium hydroxide to prepare 30% magnetic bead suspension;

[0040] Step B) surface modification of nano ...

Embodiment 2

[0043] Embodiment 2: The real-time fluorescent quantitative PCR detection of HBVDNA

[0044] The same method as in Example 1 was used to prepare magnetic beads.

[0045] Prepare the lysate and wash solution mixed with magnetic beads according to the following component ratios:

[0046] 0.3N Sodium Hydroxide, 0.45M Potassium Chloride, 0.03% Sodium N-Lauroyl Sarcosinate, 5mM EDTA, 0.45M Tris-HCL, 1% Triton X-100, 3mMDTT and 0.001wt% Bromophenol Blue.

[0047] Add the magnetic beads that have naturally precipitated for 24 hours according to the volume ratio of magnetic beads and lysate of 1:100, and mix well.

[0048] Washing liquid: 0.3M potassium chloride, 0.001wt% stone green, adjust the pH value to 4 with acetic acid.

[0049] Use the above lysate and wash solution for clinical diagnosis and the tested HBVDNA quantitative value is 5IU / ml, 10IU / ml, 20IU / ml, 2×10 2 IU / ml, 2×10 3 IU / ml, 2×10 4 IU / ml, 2×10 5 IU / ml, 2×10 6 IU / ml, 2×10 7 IU / ml, 2×10 8 IU / ml, 2×10 9 The he...

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Abstract

The invention provides a nano magnetic bead for nucleic acid purification. A preparation method of the nano magnetic bead includes the processes of preparation and surface modification of nano microspheres. The biologically-magnetic microspheres of certain sizes are obtained through an aqueous phase reaction, groups capable of adsorbing nucleic acid are introduced to the surfaces of the biologically-magnetic microspheres, through the character that silicon dioxide is electrified to adsorb nucleic acid, multiple complex samples can be efficiently purified, and therefore automation of nucleic acid purification is achieved and the workload of working staff is greatly reduced. The nano magnetic bead has the advantages that dispersibility is good, cost is low, particle uniformity is good, and the size of the magnetic bead is high in controllability, the nano magnetic bead can be effectively used for diagnostic reagent earlier purification and clinical nucleic acid detection, and has wide application prospects and social and economic benefits in the field of sequencing and the fields of biologically-magnetic microsphere targeted therapy and biological medicine development.

Description

technical field [0001] The invention relates to the field of material science, in particular to a nano magnetic bead used for nucleic acid purification. Background technique [0002] In recent years, nano-magnetic beads (abbreviated as magnetic beads) have become more and more popular, and have been widely used in many fields such as cell separation, enzyme immobilization, and nucleic acid purification. Magnetic beads refer to composite polymer microspheres with certain magnetism and special structure formed by specific combination of magnetic inorganic particles and organic polymers through physical, chemical or biological methods. The magnetic bead method is to lyse the cells first, and specifically adsorb the free nucleic acid molecules to the surface of the magnetic particles, while leaving impurities such as proteins in the solution. Then, under the action of a magnetic field, the magnetic particles are separated from the liquid, and the pure nucleic acid can be obtain...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J20/10B01J20/28C12N15/10H01F1/42
CPCB01J20/103B01J20/06B01J20/28004B01J20/28009B01J20/28019C12N15/1013H01F1/42
Inventor 王海滨王棽周其玲冯小霞
Owner BEIJING NAGENE DIAGNOSTIC REAGENT CO LTD
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