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Preparation method of contraceptive microneedle

A micro-needle and protease inhibition technology, applied in the field of immunology, can solve the problems of poor antibody specificity and poor immunogenicity, achieve good safety and effectiveness, and improve vaccine protection efficiency and bioavailability.

Inactive Publication Date: 2015-12-09
THE FIRST AFFILIATED HOSPITAL OF THIRD MILITARY MEDICAL UNIVERSITY OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In recent years, some progress has been made in the research of Eppin-based contraceptive vaccines, but there are still the following problems: (1) The use of Eppin dominant epitope vaccines for immunization can reduce adverse reactions, but the immunogenicity is poor; (2) Epitope cross-linking carriers The method of protein can enhance the immunogenicity, but the specificity of the antibody produced is poor; (3) In the experimental research of contraceptive vaccine based on Eppin, immune adjuvants such as Freund's adjuvant are usually used in combination, and its application in humans has safety problems ; Therefore, the key to solving this problem is to build a safer and more effective contraceptive vaccine delivery system
At present, there are relatively few studies on microneedle combined with immune contraception

Method used

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  • Preparation method of contraceptive microneedle
  • Preparation method of contraceptive microneedle
  • Preparation method of contraceptive microneedle

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1 Construction and Characterization of the Predominant Epitope Self-Assembled Nanocontraceptive Vaccine of Human Eppin Protein

[0047] 1. Equipment and materials

[0048] 1 device

[0049] TECNAI electron microscope (Philip company in the Netherlands); circular dichroism spectrometer (JASCO company in Japan); Model550 Microplatereader microplate reader (BIO-RAD company in the United States); S.HH.W2 digital display constant temperature water tank (Shanghai Pudong Rongfeng Scientific Instrument Co., Ltd. company); precision electronic balance (Germany Sartorius company); Vortex-Genie2 vortex oscillator (U.S. Scientific Industries company);

[0050] 2 Main experimental reagent materials

[0051] Recombinant human Eppin protein; T10022 copper mesh carbon support membrane (Beijing Xinxing Bairui Technology Co., Ltd.); uranyl acetate (SPISupplies, USA); complete Freund's adjuvant (Sigma-aldrich, USA); incomplete Freund's adjuvant (Sigma-aldrich, USA); streptavidi...

Embodiment 2

[0079] Example two cell activity

[0080] A culture of mouse bone marrow-derived dendritic cells (mBMDCs)

[0081] One female C57BL / 6 mouse was sacrificed by cervical dislocation, soaked in 75% alcohol for 10 minutes, removed the bilateral femur and tibia with ophthalmic scissors and forceps under aseptic condition, soaked in 75% alcohol, carefully removed the bone tissue, and put Soak the femur and tibia in PBS, draw 1640 culture solution with a 1mL syringe for later use, cut off the epiphysis of the long bone, insert the needle from one end of the bone into the bone marrow cavity, wash the bone marrow into a sterile petri dish, and wash each bone repeatedly 5 times until the bone turns white, pipette carefully to make bone marrow cell suspension, collect the cell suspension in the culture dish, wash the culture dish with 1640 culture medium for 3 times, collect the washing liquid, centrifuge at low temperature, 1500rpm×10min, discard Add 5mL sterile erythrocyte lysate to th...

Embodiment 3

[0095] Example 3 Preparation of microneedles

[0096] A microneedle coated with antigen

[0097] Stamp-type microneedle is a hand-held Class I medical device, mainly used for transdermal delivery of liquid drugs, which can create temporary microporous channels on the surface of the skin through the micron-scale needle tip array on its surface, thereby temporarily destroying the cutin of the skin The barrier function of the layer allows the drug to enter the deep layer of the skin. The microneedles used in this example are of two specifications: 500 μm and 1000 μm. The surface is arranged in a 6×6 array with a pitch of 500 μm, and 36 microneedle tips are evenly arranged. For details, see Figure 12 .

[0098] Transcutaneous immunization was carried out by microneedle-coated antigen method. Firstly, a precision microneedle dipping and coating device was prepared according to the array distribution and microneedle size of stamp-shaped microneedles. Using laser etching and d...

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Abstract

The invention belongs to the field of immunology, and particularly relates to development technologies of contraceptive vaccines and in particular relates to a preparation method of a contraceptive microneedle. The preparation method specifically comprises the following steps: serially connecting a sorted dominant antigen epitope sequence of a human Eppin protein and / or a self-assembled polypeptide sequence with a nanofiber structure to obtain an epitope peptide; fully dissolving the epitope peptide obtained in the step A in ultrapure water and mixing with PBS (Phosphate Buffer Solution) to obtain a self-assembled systematic working liquid; and immersing a microneedle in the self-assembled systematic working liquid for no less than 5 minutes, and taking out the microneedle and airing to obtain the contraceptive microneedle. Compared with subcutaneous injection, the contraceptive vaccine is percutaneously delivered, so that the protective efficiency and the bioavailability of the vaccine can be improved. The E-Q11 self-assembled nano vaccine has good safety and effectiveness.

Description

technical field [0001] The invention belongs to the field of immunology, and in particular relates to the development technology of contraceptive vaccines. Background technique [0002] The continuous growth of the world's population and the high incidence of unwanted pregnancy are serious problems that plague human survival, health, social economy and development. Immune contraception is considered by most researchers to meet the needs of the above-mentioned new generation of contraceptive measures, so it has become the focus of research in the field of reproduction and contraception and a contraceptive approach with great research prospects. Contraceptive vaccine (contraceptive vaccine, CV) originated from the presence of specific antibodies related to fertility suppression in immune infertility patients. The principle of immune contraception is to select key antigen components closely related to the reproductive process to make vaccines, and induce recipients to produce s...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K38/57A61K9/02A61P15/16C07K19/00A61M37/00
Inventor 梁志清唐帅何畏王延洲孙黎黎徐萍萍
Owner THE FIRST AFFILIATED HOSPITAL OF THIRD MILITARY MEDICAL UNIVERSITY OF PLA
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