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Preparation method and application of bacterial spore functional microsphere

A technology of bacterial spores and microspheres, which is applied in the field of preparation of functional microspheres of bacterial spores, which can solve the problems of human and environmental hazards, high cost, loss of activity, etc., and achieve the effects of stable mechanical rigid structure, stable and reliable performance, and simple operation

Inactive Publication Date: 2015-09-23
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the modification of the surface of polystyrene microspheres is mainly through chemical reactions (such as grafting and copolymerization) to bond specific functional groups to the surface of the microspheres. [8] , its disadvantages are: it often requires multiple complex processing steps; the chemical reagents used in the anchoring process often damage the surface structure and other properties of the particles, and there are potential hazards to the human body and the environment, etc. [3]
However, the large specific surface area and high collision frequency of nanoparticles make them have a large surface energy and tend to aggregate spontaneously, which affects the stability of nanoparticles and eventually loses their inherent activity. [11]
In addition, the process of using nanoparticles can lead to accidental spills and contamination, and is costly and inconvenient to handle

Method used

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  • Preparation method and application of bacterial spore functional microsphere
  • Preparation method and application of bacterial spore functional microsphere
  • Preparation method and application of bacterial spore functional microsphere

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Embodiment 1 (preparation embodiment)

[0044] In this example, spores of Bacillus megaterium, Bacillus subtilis and Bacillus amyloliquefaciens were used as starting materials for preparing functional microspheres (but not limited to the above materials).

[0045] The specific steps are as follows:

[0046] (1) Prepare LB liquid medium and LB solid medium, and sterilize the prepared LB liquid medium and LB solid medium at 121° C. for 30 min. Activate the above three bacillus strains with LB liquid medium for 4-6 hours, when OD 600 When = 0.6, add 200uL of activated bacterial solution to each petri dish and spread evenly, then cultivate in a 37°C incubator. The bacillus spores cultured for 3-10 days were collected, washed with deionized water and centrifuged, and repeated 2-5 times.

[0047] (2) The spores were resuspended with deionized water, and an appropriate concentration (preferably in the range of 10 8 ~10 11 spores / mL) were subpackaged in centrifuge tubes, u...

Embodiment 2( application Embodiment 1

[0051] See Example 1 for the method of preparing spore microspheres. On the basis of Example 1, the obtained biological spore microspheres are further processed, and the specific steps are as follows:

[0052] (1) Prepare the following metal nanoparticles (materials) in the laboratory: colloidal gold (particle size 15nm), colloidal silver (particle size 10-15nm), colloidal platinum (particle size 5nm), colloidal palladium (particle size 5nm), Cubic palladium (particle size 20-30nm).

[0053] (2) After mixing the prepared metal nanoparticles with the spore suspension prepared in Example 1 (the preferred concentration range is 10 8 ~10 10 per mL), placed in a constant temperature shaker for shaking incubation, so that the metal nanoparticles are adsorbed on the surface of the spores to form composite microspheres of spore metal nanoparticles.

[0054](3) The three kinds of natural spores (including but not limited to the above-mentioned spores), the processed spores, metal na...

Embodiment 3( application Embodiment 2

[0057] The spore colloidal gold composite microsphere prepared in Example 2 is used as a carrier for the detection of ApxIVA antibody in the serum of Actinobacillus pleuropneumoniae infection, and the specific application steps are as follows:

[0058] (1). Take 100 μL of microspheres and react with 200 μL of ApxIVA antigen protein to immobilize ApxIVA antigen protein on the surface of the microspheres.

[0059] (2) Bovine serum albumin (BSA) solution was used as a blocking agent to block the microspheres labeled with ApxIVA antigenic protein to occupy active sites on the surface of the microspheres that were not adsorbed by the antigenic protein to reduce non-specific reactions.

[0060] (3) Add 200 μL of the serum to be tested. If the serum contains the corresponding antibody, it can have a specific immune reaction with the antigen protein to form a microsphere-antigen-antibody complex.

[0061] (4) Add 200 μL of biotin-labeled goat anti-pig IgG to carry out the second immun...

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Abstract

The invention belongs to the field of the preparation of a biological material, and relates to a preparation method and an application of a bacterial spore functional microsphere. By virtue of a novel biochemical method, bacterial spores are processed so as to prepare a functional microsphere which is high in dispersity, and a nano material is immobilized on the surfaces of the processed pores; the functional microsphere can be applied to various fields. The preparation method is characterized by comprising the following steps: collecting bacterial spores which are cultivated for 3-10 days and are at 108-1011cfu in concentration; carrying out centrifugal washing and ultrasonic treatment; processing the bacterial pores with a trypsin liquid and a spore coating removal processing fluid, so as to finally obtain a bacterial spore microsphere that spore exosporium and spore coating are removed and surface is smooth; the microsphere is used for loading metal nanoparticles. The functional microsphere prepared by the invention is more stable in mechanical rigid structure, the loaded nano material is not easy to peel off, and the functional microsphere, as a carrier, is more stable and reliable in performance. The invention also discloses an application of the prepared spore microsphere in an animal immunization detection material.

Description

technical field [0001] The invention belongs to the technical field of biological material preparation, and in particular relates to a preparation method and application of bacterial spore functional microspheres. Background technique [0002] Functional monodisperse microspheres have extremely broad application prospects in microelectromechanical systems, optoelectronic components, biosensors, environmental analysis and catalyst carriers, etc. [1] . Polystyrene (polystyrene) is the most commonly used microsphere preparation material, usually synthesized by chemical methods, such as: latex polymerization and precipitation polymerization, etc. [2] . However, it is not an easy task to prepare microspheres with uniform shape, size, surface and optical properties using traditional mixing and extrusion techniques [3] . In addition, chemical synthesis methods inevitably use organic reagents that are harmful to humans and the environment. [0003] Modification of different act...

Claims

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Application Information

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IPC IPC(8): C12N11/14G01N33/553C12R1/11C12R1/125C12R1/07
Inventor 胡涌刚项玉强曾志明
Owner HUAZHONG AGRI UNIV
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