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Magnetic corpuscles coupled with streptavidin and biological isolation and immunoassay method

A technology of streptavidin and magnetosomes, applied in biological testing, measuring devices, analytical materials, etc., can solve the problems of extreme reaction temperature, use of toxic reagents, complex and laborious post-modification, etc., and achieve low superparamagnetism, The effect of good biocompatibility

Inactive Publication Date: 2019-01-29
湖南华腾制药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the surface of the magnetic particles in this method only has amino groups, so the cross-linking effect with streptavidin is poor, the cross-linking rate is low, and the source of the particles is chemical synthesis
Chemical synthesis of magnetic nanoparticles generally has problems such as extreme reaction temperature, difficult control of reaction conditions, use of toxic reagents, complex and laborious post-modification, etc. At the same time, its uniformity, crystallinity, purity, particle size distribution, crystal form control and operability have all far inferior to biosynthesized magnetosomes

Method used

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  • Magnetic corpuscles coupled with streptavidin and biological isolation and immunoassay method

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Experimental program
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Effect test

Embodiment 1

[0026] Magnetic Separation Column Chromatography Separation of Magnetosomes

[0027] Magnetotactic bacteria ( Magnetospirillum magneticum AMB-1) strains were inoculated in 5 L enhanced magnetic

[0028] In the spirochete culture medium, the total amount of iron quinate added was 60 μmol / L, and cultured at 26°C for 96 h. After the cultivation was completed, the fermented broth of the magnetotactic bacteria was centrifuged at 3000 r / min for 25 minutes, and then the bacteria were collected by means of a energized solenoid. The bacteria were suspended in HEPES buffer at a mass-volume ratio of 1:8. The magnetosomes are collected after crushing by a high-pressure homogenizer with a pressure of 80 MPa, and then separated by magnetic separation column chromatography.

Embodiment 2

[0030] Separation of Magnetosomes by Supercritical Fluid Extraction

[0031] Magnetotactic bacteria ( Magnetospirillum magneticum AMB-1) strains were inoculated in 5 L enhanced magnetic

[0032] In the spirochete culture medium, the total amount of iron quinate added was 60 μmol / L, and cultured at 26°C for 96 h. After the cultivation was completed, the fermented broth of the magnetotactic bacteria was centrifuged at 3000 r / min for 25 minutes, and then the bacteria were collected by means of a energized solenoid. The bacteria were suspended in HEPES buffer at a mass-volume ratio of 1:8. The magnetosomes are collected after crushing by a high-pressure homogenizer with a pressure of 80 MPa and then separated by supercritical fluid extraction. (Carry out supercritical fluid extraction with carbon dioxide, add 5% ethanol solution, extraction temperature is 50°C, extraction pressure is 13MPa, extraction time is 15min)

Embodiment 3

[0034] Separation of Magnetosomes by Subcritical Fluid Extraction

[0035] Magnetotactic bacteria ( Magnetospirillum magneticum AMB-1) strains were inoculated in 5 L enhanced magnetic

[0036] In the spirochete culture medium, the total amount of iron quinate added was 60 μmol / L, and cultured at 26°C for 96 h. After the cultivation was completed, the fermented broth of the magnetotactic bacteria was centrifuged at 3000 r / min for 25 minutes, and then the bacteria were collected by means of a energized solenoid. The bacteria were suspended in HEPES buffer at a mass-volume ratio of 1:8. Magnetosomes are collected after crushing by a high-pressure homogenizer with a pressure of 80 MPa and then extracted and separated by subcritical fluid. (Subcritical fluid extraction with butane, extraction temperature 43°C, extraction pressure 3 MPa, extraction time 30min)

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Abstract

The invention discloses magnetic corpuscles coupled with streptavidin and a biological isolation and immunoassay method. The streptavidin is cross-linked on the surfaces of the magnetic corpuscles through chemical covalent bonds, and the magnetic corpuscles have amino, carboxyl, mercapto, or hydroxyl functional groups on the surfaces of the magnetic corpuscles. The preparation process of the magnetic corpuscles includes two steps of extraction of the magnetic corpuscles and coupling of the streptavidin. The magnetic corpuscles coupled with the streptavidin has the advantages of good cross-linked effect, high biocompatibility, high magnetic controllability, super paramagnetism and low toxicity.

Description

technical field [0001] The invention belongs to the technical field of biological materials, and in particular relates to magnetosomes coupled with streptavidin and biological separation and immunodetection methods. Background technique [0002] Magnetotactic bacteria are Gram-negative bacteria that rely on the magnetic nanoparticles produced by biomineralization in vivo to move directionally under the action of an external magnetic field, and are generally microaerobic or anaerobic. The magnetic nanoparticles inside are magnetosomes, which are assembled into long chains in the magnetotactic bacteria, arranged along the long axis of the bacteria, and guide the movement of the bacteria in the external magnetic field. The magnetosome is mainly composed of an outer lipid membrane and an inner magnetite crystal. The outer membrane of the magnetosome is formed by the invagination of the magnetotactic bacterial cell membrane, and the outer membrane is negatively charged as a whole...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/53G01N33/543G01N33/569
CPCG01N33/53G01N33/54326G01N33/56911
Inventor 邓泽平罗容许慧陈瑶陈选张安林刘赛文
Owner 湖南华腾制药有限公司
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