In-vitro culture, induction, activation and cryopreservation method and cell bank establishment of immune cells

A technology of immune cells and cryopreservation method, applied in the biological field, can solve the problems of natural killer cell culture, induction, activation, cryopreservation methods need to be improved, the amplification multiple and tumor killing effect are not good, and the culture system has safety risks. , to meet the needs of a large number of immune cells, good cell viability, and low cost

Pending Publication Date: 2020-07-28
QINGDAO RESTORE BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, there are NK cell therapy products on the market, but the culture system used is complicated, the expansion multiple and tumor killing effect are not good, the recovery effect after cryopreservation is poor, and some culture systems have safety risks, etc.
Therefore, the methods of culturing, inducing, activating, and cryopreserving natural killer cells need to be improved

Method used

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  • In-vitro culture, induction, activation and cryopreservation method and cell bank establishment of immune cells
  • In-vitro culture, induction, activation and cryopreservation method and cell bank establishment of immune cells
  • In-vitro culture, induction, activation and cryopreservation method and cell bank establishment of immune cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Using the immune cell culture, induction, activation and expansion method of the present invention, the isolated mononuclear cells are induced, activated and expanded into target immune cells (NK cells), and the activity of the target immune cells is detected.

[0034] 1. Experimental method

[0035] 1. Donor Screening

[0036] 1.1 The hospital is required to sign an informed consent form with the donor in triplicate. One for the donor and one for the medical institution, and one for the laboratory along with the specimen.

[0037] 1.2 The hospital asks for the donor's personal information, past treatment history, family genetic history, and whether there is a history of infectious diseases and abnormal conditions of the hematopoietic or immune system by means of inquiries and filling in forms. The hospital is required to obtain the consent of the donor himself or his authorized personnel to check the medical examination data and obtain the medical examination informa...

Embodiment 2

[0081] The NK cells induced, activated and expanded in vitro in Example 1 were cryopreserved, and the effects of cryopreservation were compared.

[0082] The NK cells obtained on the 14th day of in vitro induction and expansion in Example 1 were cryopreserved using the following four cryopreservation solutions, and the components of the four cryopreservation solutions were as follows:

[0083] Freezing solution 1: 5% by volume of DMSO, 1% by volume of albumin, 1% by volume of aminoethanol, and 93% by volume of X-VOVO15 serum-free medium;

[0084] Freezing solution 2: 5% by volume of DMSO, 2% by volume of albumin, 2% by volume of aminoethanol, and 91% by volume of X-VOVO15 serum-free medium;

[0085] Freezing solution 3: 5% by volume of DMSO, 2% by volume of albumin, and 93% by volume of X-VOVO15 serum-free medium;

[0086] Freezing solution 4: 5% by volume of DMSO, 2% by volume of aminoethanol, 93% by volume of X-VOVO15 serum-free medium;

[0087] Freezing solution 5: 5 volu...

Embodiment 3

[0096] Using the method for culturing, inducing, and activating immune cells in the embodiment of the present invention, the isolated mononuclear cells are induced and expanded into immune cells, stored according to ABO / RH typing and HLA typing, and immune cell information available for retrieval is established Archives, Construction of immune cell banks.

[0097] 1. Donor Screening

[0098] The hospital asks for the donor's personal information, past treatment history, family genetic history, and whether there is a history of infectious diseases and abnormal conditions of the hematopoietic or immune system by means of inquiries and filling in forms. The hospital must sign an informed consent form with the donor to obtain the consent of the donor himself or his authorized personnel, to check the medical examination data and obtain the medical examination information. Donor physical examination information should include the following items: HIV-1 / 2 antibody, HBsAg, anti-HCV, ...

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Abstract

The invention discloses an in-vitro culture, induction, activation and cryopreservation method and cell bank establishment of immune cells. The method comprises the following steps of carrying out first-stage proliferation culture on mononuclear cells by using a special proliferation culture medium for the immune cells to obtain primarily proliferated immune cells; carrying out second-stage induction and proliferation culture on the primarily proliferated immune cells by utilizing a special induction culture medium for the immune cells to obtain induced immune cells; carrying out third-stage activation and proliferation culture on the induced immune cells by utilizing a special activation culture medium for the immune cells to obtain a large number of immune cells with activation functions; cryopreserving the immune cells by using a special cryopreservation solution for the immune cells to obtain cryopreserved immune cells; and performing storage according to ABO/RH typing and HLA typing, establishing an immune cell information file for retrieval, and constructing an immune cell bank.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to methods for culturing, inducing, activating and freezing immune cells in vitro and methods for establishing cell banks. Background technique [0002] Cellular biological therapy has become the fifth largest treatment mode after surgery, radiotherapy and chemotherapy, and endocrine therapy. Adoptive immune cell therapy is one of the methods of cell biological therapy. It refers to the infusion of immune cells with anti-tumor activity to tumor patients, directly killing tumor cells or stimulating the body's immune response to kill tumor cells, so as to achieve the purpose of treating tumors. Currently clinically used immune cells include DC-CIK cells, TIL cells, LAK cells and NK cells, among which CIK cells, LAK cells and NK cells are all anti-cancer systems with natural killer cells as the main body. [0003] NK cells, also known as natural killer cells, patrol the whole body of the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783C40B50/06A01N1/02
CPCC12N5/0646C40B50/06A01N1/0221C12N2501/2302C12N2501/2301C12N2501/231C12N2501/235C12N2501/14C12N2501/37C12N2533/52C12N2501/117C12N5/0031C12N2501/392C12N2501/155A01N1/0284C12N15/1037C12N2500/90C12N2501/115C12N2501/998
Inventor 张炳强陈梦梦
Owner QINGDAO RESTORE BIOTECHNOLOGY CO LTD
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