Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Cultivation system for in-vitro amplification of lymphocytes and amplification method and application

A culture system and lymphocyte technology, applied in cell culture active agents, cells modified by introducing foreign genetic material, animal cells, etc., can solve the problem of difficulty in obtaining a sufficient number of NK cells, and achieve uniform quality, strong lethality, high purity effect

Active Publication Date: 2017-09-19
杭州中赢生物医疗科技有限公司
View PDF4 Cites 7 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] One of the main obstacles restricting the clinical application of NK cells is that it is difficult to obtain a sufficient number of NK cells

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
  • Cultivation system for in-vitro amplification of lymphocytes and amplification method and application
  • Cultivation system for in-vitro amplification of lymphocytes and amplification method and application
  • Cultivation system for in-vitro amplification of lymphocytes and amplification method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] The preparation of embodiment 1 genetic engineering cell

[0057] The preparation method of the genetically engineered cells is as follows: Firstly, a vector capable of stably expressing transmembrane interleukin 21, interleukin IL12A, transmembrane interleukin 12B and CD80 was constructed. There is a selectable marker gene on the vector. Transmembrane interleukin 21 or transmembrane interleukin 12B is connected to the cell membrane through the transmembrane part of CD4, and interleukin IL12A and CD80 are transmembrane proteins. These vectors were transfected into K562 cells, and high-expressing cell clones were selected with antibiotics and flow cytometry.

Embodiment 2

[0058] The preparation of embodiment 2 lymphocyte expansion culture system

[0059] The genetically engineered cells prepared in Example 1 are used for the expansion of lymphocytes, comprising the following steps:

[0060] (1) Inactivated genetically engineered cells: irradiate with 100Gy radiation for 30 minutes to obtain inactivated K562 engineered cells;

[0061] (2) Preparation of culture medium: take lymphocyte culture medium RPMI1640 and 10% fetal bovine serum, then add the inactivated K562 engineered cells of step (1) to the mixed medium, and the addition amount is 1×10 6 / mL, then add IL-2, the amount of IL-2 added is 50U / mL.

Embodiment 3

[0062] Example 3 Expansion of Lymphocytes from Healthy Volunteers

[0063] (1) Culture: Collect blood from fresh healthy volunteers, collect serum by centrifugation for storage, and obtain human peripheral blood mononuclear cells (PBMC) through lymph separation fluid separation, and the inoculation density is 1×10 6 / mL (according to the ratio of the K562 engineered cells to the monocytes is 1:1), inoculated into the culture system of in vitro amplified lymphocytes;

[0064] (2) Amplification: 37°C, 5% CO 2 Cultivate under conditions for 7 days, then add the above culture system, and cultivate for another 7 days;

[0065] (3) Harvesting: NK cells were collected by centrifugation after the culture was completed.

[0066] NK cells co-cultured with PBMC cells from K562 cells transfected with transmembrane interleukin 21 and CD137L were used as controls;

[0067] Use an automatic cell counter to count the expanded NK cells, and draw a curve, the growth curve is as follows fig...

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
Inoculation densityaaaaaaaaaa
Login to View More

Abstract

The invention relates to the field of cellular immunotherapy, in particular to a cultivation system for in-vitro amplification of lymphocytes, an amplification method and application. The cultivation system comprises a mixture of interleukin 21, interleukin 12 and CD80. NK cells amplified through the cultivation system are high in purity and high in killing ability for tumor cells; on the 14th day, the NK cells can be amplified by about 9,100 times, so as to meet various application requirements of expected clinical treating effects on diseases; the problem on safety of the cultivation system for clinical application is solved, and the application cost of the NK cells is reduced; a foundation is laid for broad spectrum application of the NK cells to adoptive immunotherapy.

Description

technical field [0001] The invention relates to the field of cellular immunotherapy, in particular to a culture system for expanding lymphocytes in vitro, an expansion method and application. Background technique [0002] NK cells (natural killer cells, Natural Killer Cell) are also called large granular lymphocytes. And γδT cells, NKT cells act as a bridge between acquired immunity and innate immunity in form and function. On the one hand, when the body is infected or traumatized, NK cells, as defenders, quickly, extensively and specifically recognize antigens through non-peptide-MHC recognition methods, clear pathogenic microorganisms and mutant cells in a timely manner, and play the role of innate immunity. On the other hand, NK cells are considered to be partly involved in the adaptive immune response and can affect the effector functions of αβT cells and B cells. [0003] During the occurrence and development of tumors, NK cells can either directly recognize tumor cel...

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): C12N5/0783C12N5/10A61K35/17A61P35/00A61P35/02A61P31/04A61P31/12A61P31/10A61P33/00A61P1/16A61P31/20A61P31/18A61P31/14
CPCA61K35/17C12N5/0646C12N2501/2302C12N2501/2312C12N2501/2321C12N2501/51C12N2502/99
Inventor 王冶陶吴忠福彭昉刘慧显王俊俊俞英豪
Owner 杭州中赢生物医疗科技有限公司
Features
  • Generate Ideas
  • Intellectual Property
  • Life Sciences
  • Materials
  • Tech Scout
Why Patsnap Eureka
  • Unparalleled Data Quality
  • Higher Quality Content
  • 60% Fewer Hallucinations
Social media
Patsnap Eureka Blog
Learn More

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

© 2025 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com