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Antibody-modification-free magnetic bead capable of quickly capturing and detecting pathogenic bacteria and preparation method of antibody modification-free magnetic bead

An antibody modification, pathogenic bacteria technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of high cost, complex antibody modification process, difficult mass production, etc., and achieve the effect of simple production

Active Publication Date: 2020-12-08
太古宙基因科技(深圳)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the magnetic beads currently available in the market for bacterial capture are mainly antibody-modified magnetic beads. The antibody modification process is complicated and costly, making it difficult to produce in large quantities.

Method used

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  • Antibody-modification-free magnetic bead capable of quickly capturing and detecting pathogenic bacteria and preparation method of antibody modification-free magnetic bead
  • Antibody-modification-free magnetic bead capable of quickly capturing and detecting pathogenic bacteria and preparation method of antibody modification-free magnetic bead
  • Antibody-modification-free magnetic bead capable of quickly capturing and detecting pathogenic bacteria and preparation method of antibody modification-free magnetic bead

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 A method for preparing antibody-free modified magnetic beads synthesized at room temperature

[0026] 1) Dissolve 2 g of iron acetylacetonate in 50% ethanol aqueous solution, fill with nitrogen to remove the air, and ultrasonically oscillate for 10 minutes to completely dissolve it;

[0027] 2) Add 4g of sodium borohydride and 1g of polyhistidine, and stir mechanically at room temperature until the solution turns completely black;

[0028] 3) After continuing to stir for 3 hours, the resulting product collected by magnetic force was washed three times with deionized water and ethanol respectively, and stored in 10 mL of ethanol solution;

[0029] 4) Collect the generated magnetic bead product by magnetic force, and wash with deionized water and ethanol three times respectively.

[0030] 5) Resuspend the above washed particles in methanol, add 0.2g of methyl iodide, stir mechanically at room temperature (22°C), react for 3h, magnetically absorb, wash with etha...

Embodiment 2

[0031] Example 2 A method for preparing antibody-free modified magnetic beads synthesized at room temperature

[0032] 1) Dissolve 5 g of iron acetylacetonate in 50% ethanol aqueous solution, fill with nitrogen to remove the air, and ultrasonically oscillate for 15 minutes to completely dissolve it;

[0033] 2) Add 9g of sodium borohydride and 3g of polyhistidine, and stir mechanically at room temperature until the solution turns completely black;

[0034] 3) After continuing to stir for 5 hours, the resulting product collected by magnetic force was washed three times with deionized water and ethanol respectively, and stored in 100 mL ethanol solution;

[0035] 4) Collect the generated magnetic bead product by magnetic force, and wash with deionized water and ethanol three times respectively.

[0036] 5) Resuspend the above washed particles in methanol, add 0.4g of methyl iodide, stir mechanically at room temperature (25°C), react for 4h, magnetically absorb, wash with ethano...

Embodiment 3

[0037] Example 3 Method for Rapid Capture and Detection of Escherichia coli Concentration Gradient Using Antibody-Free Modified Magnetic Beads

[0038] 1) Using conventional laboratory culture methods, pick Escherichia coli colonies, inoculate, and culture at 37°C;

[0039] 2) Use sterile Tris-Ac buffer to dilute Escherichia coli colonies gradiently, and adjust the cell concentration to 10 5 cfu / ml, 10 4 cfu / ml, 10 3 cfu / ml, 10 2 cfu / ml, 10cfu / ml, take 1mL of each concentration of strains in a 1.5mL centrifuge tube, centrifuge to remove the supernatant, add 100μL PBS to the centrifuge tube, shake gently to resuspend the bacteria.

[0040] 3) Add 40ul modification-free antibody magnetic beads to the centrifuge tubes of each concentration of strains, incubate for 8 minutes, and magnetically enrich. After washing, the magnetic beads are transferred to PCR eight-tubes for RT-PCR detection, and the detection results are obtained.

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Abstract

The invention provides an antibody-modification-free magnetic bead capable of quickly capturing and detecting pathogenic bacteria and a preparation method of the antibody-modification-free magnetic bead. The magnetic bead prepared by the method is simple to prepare and the method is a one-tube in-situ two-step method without additional modification. The prepared magnetic bead quickly captures pathogenic bacteria by utilizing an electrostatic adsorption principle, and meanwhile, due to high biocompatibility of the magnetic bead, the magnetic bead carrying bacteria can be directly added into a PCR reaction solution, bacterial lysis and nucleic acid amplification are synchronously carried out, and a pathogenic bacteria detection result is simply and quickly obtained.

Description

technical field [0001] The invention relates to an antibody-free modified magnetic bead capable of rapidly capturing and detecting pathogenic bacteria and a preparation method thereof. Background technique [0002] Pathogenic bacteria affect food safety and human health. After they invade the body, they will secrete a large amount of biological toxins, thereby destroying the structure and function of the body and causing host infection. Early diagnosis of pathogenic bacteria and timely use of effective antibacterial drug treatment will help the corresponding patients recover early. Therefore, a method for early and accurate detection of pathogenic bacteria has become extremely important. Traditional bacterial detection methods require a multi-step process, which is complicated and difficult, such as how to achieve efficient capture of bacteria in complex biological samples, how to lyse bacteria after capture and achieve release of nucleic acid extraction, etc. The magneti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/686C12Q1/10C12Q1/04
CPCC12Q1/686C12Q2563/143C12Q2563/149C12Q2547/101C12Q2521/107
Inventor 戴恒
Owner 太古宙基因科技(深圳)有限公司
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