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Preparation of antigen sensitized human dendron shaped cell and its use

A technology of dendritic cells and cells, applied in the fields of biology and medicine, can solve the problem of ineffective preparation of tumor antigens

Inactive Publication Date: 2006-03-29
上海海欣生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0013] However, to date, there is no efficient method for preparing tumor antigens that elicit specific immune responses against tumors

Method used

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  • Preparation of antigen sensitized human dendron shaped cell and its use
  • Preparation of antigen sensitized human dendron shaped cell and its use
  • Preparation of antigen sensitized human dendron shaped cell and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Preparation of tumor antigens

[0077]Take surgically removed aseptic colon cancer tumor tissues (4 cases), soak them in 0.05% chlorhexidine acetate solution for 30 minutes, wash them with sterile saline three times, cut the tumor tissues into pieces with sterile tissue scissors, and add 5ml RPMI1640 medium was ground, and the single cell suspension was collected after filtering through a 200-mesh steel mesh, and then the cells were resuspended in RPMI1640 medium (1×10 7 / ml), put into a 5ml sterile cryopreservation tube, heat in a 42°C water bath for 1 hour, then continue heating in a 60°C water bath for 1 hour, quickly immerse the cryopreservation tube in liquid nitrogen for quick freezing, take it out after 10 minutes, and thaw at room temperature; Freezing and thawing was repeated 5 times in this way, the tumor cell lysate was added to a 15ml Falcon centrifuge tube, centrifuged at 400×g for 20 minutes, and the supernatant was collected. The tumor cell freeze-thawed...

Embodiment 2

[0080] Preparation of Dendritic Cells

[0081] Slowly add the collected mononuclear cells (about 50-100ml) into several 50ml Falcon centrifuge tubes pre-added with 20ml of lymphocyte separation medium (the volume of the cell suspension is equal to the volume of the separation medium). Centrifuge for 30 minutes. After centrifugation, use a flat-mouth Pasteur pipette to absorb the interfacial cells, add them to 30ml RPMI1640 medium, centrifuge at 200×g for 10 minutes, wash 3 times, and collect the cells. RPMI1640 medium with bovine serum was prepared into 4×10 6 / ml of cell suspension, seeded into a 750ml sterile Falcon culture bottle, the inoculation volume was 25ml, and the culture bottle containing the cells was placed in an incubator at 37°C, 5% carbon dioxide, and saturated humidity for 2 hours, and the culture bottle was Take it out, shake gently ten times to suspend the non-adherent cells, discard the suspended cells, then slowly add 10ml RPMI1640 medium into the bottle,...

Embodiment 3

[0084] Sensitization of dendritic cells

[0085] The harvested dendritic cells were mixed with fresh complete medium containing 10% fetal bovine serum, rhGM-CSF 50ng / ml, rhIL-410ng / ml to make 2×10 6 / ml cell suspension, add the tumor antigen prepared in Example 1 to a final concentration of 50 μg / ml, cultivate overnight, collect the dendritic cell suspension incubated overnight with the tumor antigen in a 50ml centrifuge tube, and centrifuge at 200 × g for 10 Minutes, collect the cells; then add 40ml of sterile saline, centrifuge at 200×g for 10 minutes, and wash for 3 consecutive times. The collected cells are antigen-sensitized human dendritic cells (APDC).

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Abstract

A method for sensibilizing the human dendritic cell, a process for preparing the antigen-sensitive human dendritic cell and storing it, and its application in antineoplastic immune are disclosed. Its advantages are high sensibilizing efficiency, long storage time, and high reviral rate after freeze storage.

Description

technical field [0001] The invention relates to the fields of biology and medicine, and more specifically relates to a method for preparing tumor antigens and a method for preparing antigen-sensitized human dendritic cells with the antigens prepared by the method. Background technique [0002] Dendritic cells (DC) are a kind of antigen-presenting cells with special functions in the normal human body. The activation of naive T cells is the "starter" of the body's immune response. In addition, DC can also directly or indirectly promote the proliferation and activation of B cells, regulate the humoral immune response; stimulate the activation of memory T cells to induce a second immune response; interact with NK to affect non-specific, natural immune responses, so DC can be said to be The "initiator" of the immune response in the body. DC is a key link in tumor immunity, which can absorb and process tumor antigens and mobilize the body's active and specific anti-tumor immune ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/435C12N5/08A61K39/00A61K45/00A61P35/00
Inventor 王建莉
Owner 上海海欣生物技术有限公司
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