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CN-120992923-B - Nanometer magnetic bead and preparation method thereof

CN120992923BCN 120992923 BCN120992923 BCN 120992923BCN-120992923-B

Abstract

The application relates to the technical field of cell sorting, in particular to a nano magnetic bead and a preparation method thereof. The nano magnetic beads comprise a magnetic core formed by Fe 3 O 4 and gamma-Fe 2 O 3 and a glucan layer coated outside the magnetic core, wherein the glucan layer contains polyethylene glycol. Through cladding polyethylene glycol outside the magnetic core, can make the magnetic bead in the storage process, more be difficult for taking place to agglomerate or gathering phenomenon, and then improved the save time of magnetic bead, make the efficiency of magnetic bead when selecting separately the cell higher.

Inventors

  • Xiong Fajun

Assignees

  • 纳绎生物科技(北京)有限公司

Dates

Publication Date
20260512
Application Date
20250828

Claims (6)

  1. 1. A method for preparing nano magnetic beads is characterized in that, The method comprises the following steps: preparing a solution containing Fe3+ and Fe2+, adding a proper amount of glucan, and fully mixing the components at 50-80 ℃, wherein the concentration of the Fe3+ is 0.01-0.05mol/l, and the concentration of the Fe2+ is 0.03-0.06mol/l; adjusting the pH of the solution to be alkaline, adding a proper amount of polyethylene glycol 600, controlling the temperature to be 50-80 ℃ for continuous reaction, and rapidly cooling the solution after the reaction is finished to obtain dextran-coated magnetic beads; placing the magnetic beads in a preservation solution to form a magnetic bead suspension of about 10 mg/ml; The preservation solution comprises the following components: 2-8% of trehalose, 2000.5-2% of polyethylene glycol, 0.05-0.1% of sodium ascorbate, 0.2-1% of BSA and 0.02-0.1% of poloxamer; EDTA 0.4-0.6mM, sodium citrate 4-6mM; The pH of the solution is 7.5-9; taking the dextran coated magnetic beads, and washing the magnetic beads with buffer solution to form magnetic bead suspension; 10mg/ml EDC and 10mg/ml NHS are added, and carboxyl groups are activated for 0.5-2h at 36-38 ℃; removing the supernatant and re-suspending with buffer solution; adding antibody, and culturing at 36-38deg.C for 6-12 hr.
  2. 2. The method for preparing the nano magnetic beads according to claim 1, wherein: The magnetic beads are placed in the preservation solution for no more than 20 days.
  3. 3. The method for preparing the nano magnetic beads according to claim 1, wherein: the antibody is a CD4 antibody.
  4. 4. The method for preparing the nano magnetic beads according to claim 1, wherein: the prepared nano magnetic beads comprise a magnetic core formed by Fe3O4 and gamma-Fe 2O3 and a glucan layer coated outside the magnetic core; Wherein the dextran layer comprises polyethylene glycol.
  5. 5. The method for preparing the nano magnetic beads according to claim 4, wherein: The polyethylene glycol is polyethylene glycol 600.
  6. 6. The method for preparing the nano magnetic beads according to claim 4, wherein: The glucan is carboxylated glucan.

Description

Nanometer magnetic bead and preparation method thereof Technical Field The application relates to the technical field of cell sorting, in particular to a nano magnetic bead and a preparation method thereof. Background Magnetic beads have been developed gradually as an important means for sorting cells due to their simplicity, high efficiency, high specificity and the like. In general, magnetic separation is mainly based on modification of antibodies, proteins and the like, and the magnetic beads are modified, and the target cells are rapidly separated under the action of an external magnetic field through the specific combination of the modification and the cell surface molecules. In the prior art, the synthesis route of the magnetic beads is generally that functionalized magnetic cores are synthesized first, and then the magnetic cores are coated into the magnetic beads and then coupled with biological ligands, so that the specific treatment of the magnetic beads is realized. However, as the particle size of the magnetic beads is reduced, the agglomeration effect is more obvious, and the specific treatment efficiency of the magnetic beads is affected, so that the separation effect on target cells is also affected. In view of this, the present application has been proposed. Disclosure of Invention The application aims to provide a nano magnetic bead and a preparation method thereof, which aim to solve at least one technical problem in the background art. In particular, in a first aspect of the application, there is provided a nanomagnetic bead, The nanometer magnetic beads comprise a magnetic core formed by Fe 3O4 and gamma-Fe 2O3 and a glucan layer coated outside the magnetic core; Wherein the dextran layer comprises polyethylene glycol. By adopting the technical scheme, polyethylene glycol is coated outside the magnetic core, so that the magnetic beads are not easy to agglomerate or aggregate in the storage process, the storage time of the magnetic beads is further prolonged, and the efficiency of the magnetic beads in cell sorting is higher. Preferably, the polyethylene glycol is polyethylene glycol 600. Preferably, the dextran is carboxylated dextran. Preferably, the nanomagnetic beads further comprise an antibody linked to the carboxylated dextran, the antibody being a CD4 antibody. In a second aspect of the present application, there is provided a method for preparing a nano magnetic bead, comprising the steps of: Preparing a solution containing Fe 3+、Fe2+, adding a proper amount of glucan, and fully mixing the components at 50-80 ℃, wherein the concentration of Fe 3+ is 0.01-0.05mol/l, and the concentration of Fe 2+ is 0.03-0.06mol/l; And (3) regulating the pH value of the solution to be alkaline, adding a proper amount of polyethylene glycol 600, controlling the temperature to be 50-80 ℃ for continuous reaction, and rapidly cooling the solution after the reaction is finished to obtain the dextran-coated magnetic beads. Preferably, the preparation method of the nano magnetic beads further comprises the steps of: The beads were placed in a preservation solution to form a suspension of about 10mg/ml of beads. Preferably, the preserving fluid comprises the following components: 2-8% of trehalose, 2000.5-2% of polyethylene glycol, 0.05-0.1% of sodium ascorbate, 0.2-1% of BSA and 0.02-0.1% of poloxamer; EDTA 0.4-0.6mM, sodium citrate 4-6mM; the pH of the solution is 7.5-9. Preferably, the magnetic beads are placed in the preservation solution for no more than 20 days. Preferably, the preparation method of the nano magnetic beads further comprises the step of connecting antibodies: taking the dextran coated magnetic beads, and washing the magnetic beads with buffer solution to form magnetic bead suspension; 10mg/ml EDC and 10mg/ml NHS are added, and carboxyl groups are activated for 0.5-2h at 36-38 ℃; Re-suspending with buffer after removing supernatant; adding antibody, and culturing at 36-38deg.C for 6-12 hr. Preferably, the antibody is a CD4 antibody. In summary, the application has the following beneficial effects: Firstly, according to the nano magnetic beads provided by the application, polyethylene glycol is coated outside the magnetic cores, so that aggregation or aggregation phenomenon of the magnetic beads is less likely to occur in the storage process, the storage time of the magnetic beads is further prolonged, and the efficiency of the magnetic beads in cell sorting is higher. Secondly, the preparation method of the nano magnetic beads provided by the application can be better matched with the magnetic bead preservation solution provided by the application, so that the preservation efficiency of the magnetic beads is improved, and the occurrence of agglomeration phenomenon is reduced. Thirdly, the nano magnetic bead preparation method provided by the application has higher binding efficiency with the antibody, especially the CD4 antibody, so that the magnetic bead has stronger specificity when