Polypeptide preparations

A peptide and amino acid technology, applied in the direction of peptides, peptide sources, peptides, etc., can solve problems such as infection or disease

Pending Publication Date: 2021-12-21
QINGDAO MARINE BIOPHARMACEUTICAL RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since CoViD-19 is a relatively novel variant, it will take a long time for our immune system to recognize and respond to CoViD-19. If the immune system develops into a serious infection or disease state before it is activated, it will can lead to fatal results

Method used

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  • Polypeptide preparations
  • Polypeptide preparations
  • Polypeptide preparations

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1: Peptide-HLA-I complex formation

[0035] The experiment was carried out according to the Monomer / Tetramer production protocol experimental operation flow of the easYmer HLA-A*02:01 MHC Tetramers Kit kit of the immunoware manufacturer.

[0036] 1. Weigh an appropriate amount of peptide to be tested (see Table 1), and dissolve it in dimethyl sulfoxide to 1 mM.

[0037] 2. The positive control peptide (the positive control peptide is provided by the immunoware easYmer HLA-A*02:01 MHCTetramers Kit), and the peptide to be tested is diluted to 25 μM with double distilled water.

[0038] 3. Prepare each reaction tube on ice according to the following table (see Table 2), and mix well by repeatedly pipetting with a pipette to avoid the generation of air bubbles.

[0039] 4. After sealing with the sealing tape, incubate at 18 degrees for 48 hours before use.

[0040] no size sequence no size sequence 1# 1,380 GVAPGTAVLRQW 19# 1,380 RTV...

Embodiment 2

[0044] Example 2: Detection of the Formation of Peptide-HLA-I Complex Based on Flow Cytometry

[0045] 1. Add 4 μl of peptide-HLA-I complex solution to 46 μl of dilution buffer and mix well.

[0046] 2. Add 25 μl peptide-HLA-I complex dilution to 50 μl dilution buffer and mix well.

[0047] 3. Transfer 40 μl of the dilution from step 2 to a new EP tube.

[0048] 4. Streptavidin-coated magnetic beads (6-8 μm), diluted 45 times with dilution buffer, added 20 μl to each tube, and mixed well.

[0049] 5. Incubate on a shaker at 37 degrees for 1 hour.

[0050] 6. Add 160 μl FACS solution to each tube.

[0051] 7. Centrifuge at 700g for 3 minutes and discard the supernatant.

[0052] 8. Resuspend in 200μl FACS solution, centrifuge at 700g for 3min, discard the supernatant, repeat this step, and wash twice.

[0053] 9. Dilute PE-labeled anti-human monoclonal antibody BBM.1 200 times with FACS solution.

[0054] 10. Add 50 μl of antibody diluent to each tube to resuspend, and in...

Embodiment 3

[0059] Example 3: Tetramer related:

[0060] This experiment was carried out in accordance with the Monomer / Tetramer production protocol experimental procedure of the easYmer HLA-A*02:01 MHC Tetramers Kit kit from the immunoware manufacturer.

[0061] Preparation of tetramers.

[0062] 1. Take 6 μL of the prepared 500nM polypeptide 18#, 23#, 30# and HLA-I monomer complex in a 0.2mL centrifuge tube, add 0.48μL Streptavidin-BV421 (BD; Cat#563259; 0.1mg / ml ).

[0063] 2. After mixing evenly, incubate at 4°C in the dark for 1 hour.

[0064] Isolation of Human Peripheral Blood Mononuclear Cells (PBMC)

[0065] 1. Take human venous anticoagulated whole blood (EDTA) and dilute it with an equal volume of normal saline.

[0066] 2. Add an appropriate amount of lymphocyte separation medium into a 15mL centrifuge tube, spread the diluted blood on top of the separation medium, and keep the interface between the two liquid surfaces clear.

[0067] 3. At room temperature, centrifuge ho...

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Abstract

On the basis of AI-Deep-Learning technical research, activation of an MHC-I immune system is taken as a target, research is carried out aiming at CoViD-19 virus structural protein, polypeptide sequences with the length of 9-14 amino acids are predicted, the polypeptides can obviously activate the immune system of a body, and a foundation is laid for designing a safe and effective polypeptide vaccine.

Description

technical field [0001] The present invention relates to biological products, especially polypeptide products. Background technique [0002] Activated T lymphocytes play an important role in anti-tumor, anti-infection, and autoimmune diseases. The activation of T cells requires two different signals. The first signal comes from the interaction between TCR (Tcell receptor, T cell receptor) and antigen peptide-MHC (major histocompatibility complex, major histocompatibility complex) complex, triggering a series of antigen-specific T cell responses, including intracellular The activation of two signaling pathways, calcineurin, MAPK and NF-κB, leads to downstream molecular events to initiate the expression of various genes, including IL-2, IL-2L receptor (CD25), CD69 and CD154, etc. The second signal comes from the co-stimulatory signal generated by the combination of B7 family molecules on APC (antigen presenting cells) and its ligand CD28 family molecules on T cells, such as t...

Claims

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Application Information

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IPC IPC(8): C07K14/165A61K39/215A61K45/06A61P37/04A61P31/12A61P31/14A61P31/16A61P31/20A61P31/22A61P31/18G01N33/569
CPCC07K14/005A61K39/12A61K45/06A61P37/04A61P31/12A61P31/14A61P31/20A61P31/22A61P31/18A61P31/16G01N33/56983C12N2770/20022C12N2770/20034G01N2333/165
Inventor 杨金波申載敏刘珊冉祥宋巧玲郑英柱姚丹杨萌琳赵晨阳
Owner QINGDAO MARINE BIOPHARMACEUTICAL RES INST
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