A preparation method of nano magnetic beads for purifying histidine-tagged proteins

A technology of nano-magnetic beads and histidine tags, which is applied in the field of preparation of nano-magnetic beads, can solve the problems of numerous and complicated steps in the synthesis process, difficult to connect to the surface of magnetic beads, and high requirements for sample loading, and achieve simple purification and separation. , low cost, short time-consuming effect

Active Publication Date: 2021-08-17
南京青柠生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this column purification method has several disadvantages: 1. The requirements for the sample solution are relatively high, and cell debris and impurities must be removed before loading the sample to avoid clogging the column
However, this patented synthesis process has many and complicated steps, and the reaction efficiency of NTA is low (acrylic acid is difficult to react with NTA under the action of EDC and NHS), and it is difficult to connect to the surface of magnetic beads. In addition, the whole process requires many reactants and is expensive , if EDC, NHS, NTA, etc., the reaction cost is high

Method used

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  • A preparation method of nano magnetic beads for purifying histidine-tagged proteins
  • A preparation method of nano magnetic beads for purifying histidine-tagged proteins
  • A preparation method of nano magnetic beads for purifying histidine-tagged proteins

Examples

Experimental program
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Effect test

Embodiment 1

[0041] This example relates to the preparation and application of a new type of nano-magnetic beads for purifying histidine-tagged proteins.

[0042] The preparation method of magnetic beads comprises the following steps:

[0043] The first step, reactant 0.5 gram sodium alginate, 0.5 gram FeCl 3 ·6H 2 O, 1.5 g NaAc 3H 2 O. 1 milliliter of PEG (MW: 300-6000) and 1 milliliter of deionized water were mixed in 20 milliliters of ethylene glycol solvent, and mechanically stirred at 300 rpm until completely dissolved. Under the conditions of reaction for 25 hours, after cooling to room temperature, the reaction product was moved to a 50 ml centrifuge tube, washed with deionized water, collected with a magnet, and stored in vacuum to obtain black Fe 3 o 4 nanoparticles.

[0044] In the second step, cross-linked sodium alginate is coated on the surface of nanoparticles: 1 g of Fe 3 o 4 Nanoparticles, 1.2 grams of sodium alginate, 0.5 grams of sodium hydroxide, 2 milliliters of ...

Embodiment 2

[0057] This example relates to the preparation and application of a new type of nano-magnetic beads for purifying histidine-tagged proteins.

[0058] The preparation method of magnetic beads comprises the following steps:

[0059] The first step, reactant 0.1 gram sodium alginate, 0.05 gram FeCl 3 ·6H 2 O, 0.05 g NaAc 3H 2 O. 0.1 milliliters of PEG (MW: 300-6000) and 0.1 milliliters of deionized water were mixed in 20 milliliters of ethylene glycol solvents, mechanically stirred at 300 rpm until completely dissolved, then the above solution was transferred to a 50 milliliter reaction kettle, and heated at 100°C Under the conditions of reaction for 48 hours, after cooling to room temperature, the reaction product was moved to a 50 ml centrifuge tube, washed with deionized water, collected with a magnet, and stored in vacuum to obtain black Fe 3 o 4 nanoparticles.

[0060] In the second step, cross-linked sodium alginate is coated on the surface of nanoparticles: 0.5 g of F...

Embodiment 3

[0066] This example relates to the preparation and application of a new type of nano-magnetic beads for purifying histidine-tagged proteins.

[0067] The preparation method of magnetic beads comprises the following steps:

[0068] In the first step, reactant 1 gram of sodium alginate, 1 gram of FeCl 3 ·6H 2 O, 3 g NaAc 3H 2 O. 2 milliliters of PEG (MW: 300~6000) and 2 milliliters of deionized water are mixed in 20 milliliters of ethylene glycol solvents, and after mechanical stirring at 300 rpm until completely dissolved, the above-mentioned solution is transferred to a 50 milliliter reaction kettle and heated at 250° C. Under the conditions of reaction for 2 hours, after cooling to room temperature, the reaction product was transferred to a 50 ml centrifuge tube, washed with deionized water, collected with a magnet, and stored in vacuum to obtain black Fe 3 o 4 nanoparticles.

[0069] In the second step, cross-linked sodium alginate is coated on the surface of nanopartic...

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Abstract

The invention discloses a preparation method of nano-magnetic beads for purifying histidine-labeled protein; 3 ·6H 2 O,NaAc·3H 2 O and PEG are used as raw materials, ethylene glycol and deionized water are used as solvents, and monodispersed sodium alginate / Fe is generated by solvothermal reaction. 3 O 4 Magnetic nano-magnetic beads, then the nano-magnetic beads are further cross-linked and activated with the newly added sodium alginate through epichlorohydrin, and then iminodiacetic acid (IDA) chelating agent is introduced to obtain efficient chelating with metal nickel or cobalt ions. Combined with water-stable magnetic nano-magnetic beads, efficient, fast, simple, and inexpensive separation and purification of histidine-tagged proteins can be achieved. The invention is simple, fast, stable and low in cost, the synthesized nano-magnetic beads have a large specific surface area, are stable in the aqueous phase and are not easy to agglomerate and precipitate, greatly improve the efficiency of purifying histidine-tagged proteins, and can be used to realize large-scale industrial production .

Description

technical field [0001] The invention relates to the technical field of biological materials, in particular to a method for preparing nano magnetic beads used for purifying histidine-tagged proteins. Background technique [0002] At present, the purification methods of histidine-tagged proteins mainly use agarose or dextran microspheres as solid phase carriers, and immobilize the chelating agent iminodiacetic acid (IDA) or nitrilotriacetic acid (NTA) on the microspheres through chemical cross-linking. surface, and then fill the column with microspheres to purify histidine-tagged proteins by means of an affinity chromatography column. However, this column purification method has several defects: 1. The requirements for the sample solution are relatively high, and cell debris and impurities must be removed before sample loading to avoid clogging the column. 2. It is necessary to control the sample loading speed to prevent the column from collapsing due to excessive pressure. ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01J13/14C08L5/04C08K3/22
Inventor 乔秀华
Owner 南京青柠生物科技有限公司
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