Self-assembly nanometer adjuvant and preparing method and application of nanometer vaccine formed by self-assembly nanometer adjuvant

A nano-vaccine and self-assembly technology, which is applied in the preparation of nano-vaccine and in the field of nano-vaccine, can solve problems such as application limitations, achieve good delivery effect, enhance bioavailability, and maintain strong immune response

Active Publication Date: 2019-06-28
北京康曜生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

CPG ODN is a promising immune adjuvant, but its application is limited due to its easy aggregation and degradation in the body

Method used

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  • Self-assembly nanometer adjuvant and preparing method and application of nanometer vaccine formed by self-assembly nanometer adjuvant
  • Self-assembly nanometer adjuvant and preparing method and application of nanometer vaccine formed by self-assembly nanometer adjuvant
  • Self-assembly nanometer adjuvant and preparing method and application of nanometer vaccine formed by self-assembly nanometer adjuvant

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] This example is the preparation process of EV71 nano vaccine formed by encapsulating EV71 inactivated virus with nano adjuvant disclosed in the present invention.

[0043](a) the preparation method comprises the following steps:

[0044] (1) Prepare 2 mg / mL carboxymethyl dextran solution with deionized water, and pass through a 0.22 μm microporous membrane;

[0045] (2) Prepare a 1 mg / mL protamine sulfate solution with deionized water, and pass through a 0.22 μm microporous membrane;

[0046] (3) dissolving the EV71 inactivated virus and CpG oligodeoxynucleotides in the carboxymethyl dextran solution in the above (1);

[0047] (4) Add the protamine sulfate solution in the above (2) dropwise to the solution in the above (3), finally protamine sulfate: carboxymethyl dextran: EV71 inactivated virus: CpG The mass ratio of oligodeoxynucleotides is 1.2:2:0.075:0.0375. Use a magnetic stirrer to continuously stir for 30 minutes, place it stably, centrifuge, remove the superna...

Embodiment 2

[0053] (a) EV71 inactivated virus group (abbreviated as ENP), EV71 inactivated virus and CpG oligodeoxynucleotide mixed group (abbreviated as EC) with EV71 nano-vaccine (abbreviated as ECNP) prepared in Example 1 and nano self-assembly carrier , EV71 inactivated virus and CpG oligodeoxynucleotide mixed group with incomplete Freund's adjuvant (abbreviated as ECFA) and PBS solution group were used as negative control (abbreviated as PBS) to immunize mice respectively, and the immunization scheme was as follows:

[0054] BALB / c female mice aged 6-8 weeks were selected as immunization objects, and the experimental animals were divided into 5 groups, 8 each in the ECNP group, ENP group, EC adjuvant group, EC group, and PBS group.

[0055] (1) For the first immunization (week 0), the injection volume of each injection is 100 μL. Injected subcutaneously.

[0056] (2) Booster immunization (2 weeks, 4 weeks), the injection volume of each needle is 100 μL, and the concentration is half...

Embodiment 3

[0076] (a) get in embodiment 2, immunize the mouse serum after 6 weeks, measure the content of IFN-α in the serum by ELISA method, concrete steps are as follows:

[0077] (1) Adding samples: Set standard wells, sample wells to be tested and blank wells respectively. Set standard wells to 7 wells, and add 100 μL of standard products with different concentrations in turn. Add 100 μL to the blank well, add 100 μL of the sample to be tested to the remaining well, add a film to the enzyme plate, and incubate at room temperature at 37 degrees for 2 hours.

[0078] (2) Discard the liquid, spin dry without washing.

[0079] (3) Add 100 μL of detection solution A working solution to each well, cover the enzyme plate with a film, and incubate at room temperature at 37 degrees for 1 hour.

[0080] (4) Discard the liquid in the wells, wash each well with 350 μL of washing solution, soak for 1-2 minutes, absorb (do not touch the plate wall) or shake off the liquid in the microplate, and ...

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Abstract

The invention relates to a self-assembly nanometer adjuvant and a preparing method and application of nanometer vaccine formed by the self-assembly nanometer adjuvant. According to the self-assembly nanometer adjuvant and the preparing method and application of the nanometer vaccine formed by the self-assembly nanometer adjuvant, protamine sulfate and carboxymethyl dextran serve as self-assembly materials, the protamine sulfate, the carboxymethyl dextran and CpG oligodeoxynucleotide form the nanometer adjuvant in a self-assembly manner, and the nanometer vaccine is formed by the nanometer adjuvant and a viral antigen. Through the nanometer adjuvant, the bioavailability of the CpG oligodeoxynucleotide is improved, it is avoided that the CpG oligodeoxynucleotide is degraded in the body, andfunctions of A-type CpG are increased through B-type CpG. Through the nanometer vaccine, a TH1-type humoral immunity response can be induced, and meanwhile a high cellular immune response can be induced. Mice in-vivo challenge experiments verify that the nanometer vaccine has a good protection effect, and provides help for future nanometer-vaccine application.

Description

technical field [0001] The invention belongs to the field of nanometer material biology. The invention relates to a nano-adjuvant formed by self-assembly and a nano-vaccine formed by the adjuvant, in particular to using protamine sulfate and carboxymethyl dextran as self-assembly materials together with CpG oligodeoxynucleotides A nano-adjuvant formed by self-assembly and a preparation method and application of a nano-vaccine formed by the nano-adjuvant and a virus antigen. Background technique [0002] Viral vaccines include live attenuated vaccines, inactivated vaccines, and subunit vaccines. The inactivated virus is composed of a complete virus, which artificially loses or weakens its pathogenicity, but still maintains all or part of the immunogenicity of the virus. After vaccination, the virus antigen can stimulate the body to produce an immune response and achieve a protective effect. Due to its poor immune effect, inability to induce CTL response, short duration of t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/39A61P37/04A61K39/125A61P31/14
CPCA61K39/12A61K39/39A61P31/14A61P37/04A61K2039/55583A61K2039/55516A61K2039/55511A61K2039/5252C12N2770/32334
Inventor 盛望梁皓邓雄威肖向茜沈思嗣
Owner 北京康曜生物科技有限公司
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