B cell vaccine based on Hepal-6 hepatoma cell autophagosome-DRibbles and preparation method of B cell vaccine

A technology of hepatoma cells and autophagosomes, which can be applied to tumors/cancer cells, animal cells, vertebrate cells, etc., can solve the problem of insufficient strength, and achieve the effect of reducing harm and improving immune response ability.

Active Publication Date: 2013-12-25
SOUTHEAST UNIV
View PDF3 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Insufficient immune response elicited by va

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • B cell vaccine based on Hepal-6 hepatoma cell autophagosome-DRibbles and preparation method of B cell vaccine
  • B cell vaccine based on Hepal-6 hepatoma cell autophagosome-DRibbles and preparation method of B cell vaccine
  • B cell vaccine based on Hepal-6 hepatoma cell autophagosome-DRibbles and preparation method of B cell vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Example 1: Electron Microscopic Observation of the Morphology of Autophagosomes-DRibbles in Hepa1-6 Liver Cancer Cells

[0059] The extracted Hepa1-6 liver cancer cell autophagosome-DRibbles was centrifuged at 12000rpm at a high speed to precipitate and compress tightly, discard the supernatant, and Hepa1-6 liver cancer cell autophagosome-DRibbles was precipitated with 2.5% (v / v) pentadiene Aldehyde was fixed, processed in the electron microscope room, and the morphology of autophagosomes-DRibbles of Hepa1-6 liver cancer cells was observed on the microscope.

[0060] Such as figure 1 Shown: Under the electron microscope, the autophagosomes of Hepa1-6 liver cancer cells-DRibbles are small bodies with a double-layer membrane structure, with an average diameter of about 300nm-1μm (pointed by the arrow), which proves that the autophagosomes of liver cancer cells are effectively recruited .

Embodiment 2

[0061] Example 2: Hepa1-6 liver cancer cell autophagosomes-DRibbles induce B cell proliferation and secretion of IgM antibodies in vitro

[0062] 1. Hepa1-6 liver cancer cell autophagosomes-DRibbles induce B cell proliferation and secretion of IgM antibodies in vitro

[0063] ①Hepa1-6 liver cancer cell autophagosomes-DRibbles stimulate B cell proliferation experiment

[0064] Take C57 / BL6 mouse splenocytes, use CD43 negative selection magnetic beads to separate mouse spleen B cells, and mark B cells with 5uM CFSE. After that, CFSE-labeled B cells were stimulated with 10ug / ml Hepa1-6 liver cancer cell autophagosome-DRibbles, Hep1-6 cell lysate and 10ug / ml LPS respectively. After 5 days, B cells were collected by centrifugation at 1000rpm, and CFSE was identified by flow cytometry + The division and proportion of B cells.

[0065] Such as figure 2 As shown, the B cells stimulated by the autophagosome-DRibbles of Hepa1-6 liver cancer cells can promote 31.6% of B cells to div...

Embodiment 3

[0069] Example 3: Hepa1-6 liver cancer cell autophagosomes-DRibbles induce B cells to produce specific antibodies in vivo

[0070] Inject C57 / BL6 mice (30ug total protein / mouse) via tail vein with Hepa1-6 liver cancer cell autophagosomes-DRibbles on the 1st, 2nd, and 3rd day, collect blood from orbit on the 7th day, separate serum (frozen at -20°C) save).

[0071] 1. Detection of IgM and IgG in serum (ELISA method)

[0072] ① Coat the 96-well plate with anti-mouse IgM or IgG (1:10000) and leave overnight at 4°C.

[0073] ② Wash the plate 3 times with PBST (phosphate buffered saline PBS containing 0.5% Tween), 3 minutes each time. Then block with 2% goat serum blocking solution, 37°C, 2h.

[0074] ③ Wash the plate 3 times with PBST. Serum samples were diluted (1:500, 1:1000, 1:2000...), added to a sealed 96-well plate, and incubated at 37°C for 1 hour.

[0075] ④ Wash the plate 5 times with PBST, add HRP-antimouse-IgM or IgG (1:10000), and incubate at 37°C for 1h.

[0076...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

PropertyMeasurementUnit
The average diameteraaaaaaaaaa
Login to view more

Abstract

The invention discloses a B cell vaccine based on Hepal-6 hepatoma cell autophagosome-DRibbles and a preparation method of the B cell vaccine. The invention firstly finds that the Hepal-6 hepatoma cell autophagosome-DRibbles used as a novel antigen carrier with an adjuvant function can be used for activating a B lymphocyte to generate a specific antibody and effectively present an antigen, and an immune mouse can induce a specific cell and humoral immune response. In addition, the Hepal-6 hepatoma cell autophagosome-DRibbles are firstly used for inducing a B cell to generate a tumor specificity antibody and effectively present a tumor antigen. The Hepal-6 hepatoma cell autophagosome-DRibbles are natural exocytosis components of a cell, and the vaccine based on the Hepal-6 hepatoma cell autophagosome-DRibbles can be used for reducing the possible harms to an organism on the basis of improving the immune response capacity, so that the vaccine is a novel antigen carrier with broad prospects.

Description

technical field [0001] The invention belongs to the technical field of tumor vaccine development, and in particular relates to a B cell vaccine based on Hepa1-6 liver cancer cell autophagosomes-DRibbles and a preparation method thereof. Background technique [0002] Autophagy is a ubiquitous phenomenon in cell biology, which refers to the encapsulation of cytoplasmic components and organelles to form autophagosomes, which then fuse with lysosomes to form autolysosomes. The process of degrading intracellular substances. [0003] A large number of research results have shown that the use of autophagy inhibitors (3-AM, wortmannin) and siRNA technology to down-regulate autophagy-related genes (Atg12) and other methods to inhibit autophagy of tumor cells can significantly reduce DC-based dendritic cell cross-presentation of tumor cells. Antigen immune effect; on the contrary, the use of autophagy inducers (rapamycin, NH 4 CL), starvation and other methods induce tumor cells to ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): A61K39/00A61K39/39C12N5/09C12N5/0781A61P35/00
Inventor 王立新曹萌
Owner SOUTHEAST UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap