Immune cell and preparation method thereof

A technology of immune cells and cells, applied in the biological field, can solve the problems of insignificant effect, low content of tumor antigen substances, and low characteristics of tumor antigens, and achieve high affinity, strong tumor antigen sensitization, and strong antigen sensitization The effect of action

Active Publication Date: 2015-11-11
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the current DC-CIK cell preparation method, the content of tumor antigens contained in the plasma obtained by centrifugation is very low, and the method of low temperature treatment and rewarming to remove the supernatant is used to concentrate the antigen components, the effect is not obvious, and then inactivation treatment , the tumor antigen characteristics are extremely low, so adding it to DC cell culture medium to stimulate DC development has poor effect

Method used

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  • Immune cell and preparation method thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0079] Example 1 Preparation of immune cells

[0080] (1) Culture of tumor cells and preparation of preparation 1

[0081] a, Tumor tissue separation Tumor cell A431 (adhesive growth tumor cell), 1640 medium+5% serum culture, inoculation density 5-10×10 5 cell / mL was inoculated in a T75 culture flask.

[0082] b. Change the liquid at half the amount the next day. The specific operation steps are as follows:

[0083] After the adherent growth tumor cells were 80% confluent, they were digested with 0.25% trypsin for 1-2 minutes, the serum was terminated, centrifuged at 400g for 5 minutes to pellet the cells, and subcultured at 1:2. Change the medium according to the cell growth status (change the medium once every 2-3 days), and collect the supernatant. After culturing for 4 days, all supernatants were collected, digested with 0.25% trypsin for 1-2 min, serum was terminated, centrifuged at 400 g for 5 min to collect cells.

[0084] c. The supernatant is concentrated by an ul...

Embodiment 2

[0102] Example 2 Preparation of immune cells

[0103] (1) Culture of tumor cells and preparation of preparation 1

[0104] a, Tumor tissue separation Tumor cells HL60 (suspension growth tumor cells), 1640 medium + 5% serum culture, inoculation density 5-10 × 10 5 cell / mL was inoculated in a T75 culture flask.

[0105] b. Change the liquid at half the amount the next day. The specific operation steps are as follows:

[0106] Rehydrate once every 2-3 days to maintain the cell density at 1-1.5×10 6 cell / mL, after 4-8 days, collect the cell suspension, centrifuge at 200-400g for 5min, and collect the supernatant and cells respectively.

[0107] c. The supernatant is concentrated by an ultrafiltration system, and the instrument uses a Sartorius Slice200 tangential flow device. Steps:

[0108] 1) The culture solution is pre-filtered through a filter membrane with a pore size of 0.45 μm to remove bulky impurities;

[0109] 2) Filter again through a filter membrane with a pore si...

Embodiment 3

[0124] Example 3 Preparation of immune cells

[0125] (1) Culture of tumor cells and preparation of preparation 1

[0126] a, Tumor tissue separation Tumor cell K562 (adhesive growth tumor cell), 1640 medium+5% serum culture, inoculation density 5-10×10 5 cell / mL was inoculated in a T75 culture flask.

[0127] b. Change the liquid at half the amount the next day. The specific operation steps are as follows:

[0128] After 90% of the adherent tumor-like cells were confluent, they were digested with 0.25% trypsin for 1-2 minutes, and the digestion was terminated with serum, centrifuged at 400g for 5 minutes to pellet the cells, and then subcultured at 1:2. Change the medium according to the cell growth status (change the medium once every 2-3 days), and collect the supernatant. After culturing for 8 days, all the supernatants were collected, digested with 0.25% trypsin for 1-2 min, the digestion was terminated by serum, and the cells were collected by centrifugation at 200 g ...

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Abstract

The invention relates to the technical field of biology, in particular to an immune cell and a preparation method thereof. The preparation method comprises the steps that tumor cell nucleotide is mixed with supernate, and a first preparation is obtained; PBMC is taken for culture, and a suspension cell and an adherent cell are separated; the adherent cell is cultured through a culture medium containing the first preparation, and a DC cell is obtained; induced culture is conducted on the suspension cell, and an immune cell with killing efficiency is obtained; the DC cell and the immune cell with the killing efficiency are cultured together, and the immune cell is prepared. According to the the immune cell and the preparation method thereof, the tumor cell nucleotide and protein allergic sources are utilized simultaneously, strengthening stimulation is conducted on the DC cell, stronger antigen sensitization is exerted, higher affinity with a tumor cell is achieved, and the risk factor of the tumor cell does not exist.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an immune cell and a preparation method thereof. Background technique [0002] Tumor is a new organism formed by the body under the action of various tumorigenic factors, and the cells of local tissues lose their normal regulation of their growth at the gene level, resulting in abnormal proliferation and differentiation. Once a new organism is formed, it does not stop growing due to the elimination of the cause. Its growth is not regulated by normal body physiology, but destroys normal tissues and organs. This is especially obvious in malignant tumors. Compared with benign tumors, malignant tumors grow faster and show invasive growth, are prone to bleeding, necrosis, ulcers, etc., and often have distant metastasis, resulting in emaciation, weakness, anemia, loss of appetite, fever, and severe organ damage. Impairment of function, etc., eventually lead to death of the patient. [00...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
Inventor 陈海佳王一飞葛啸虎应杰
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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