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BsAb in-vitro loaded T cell

A technology of external loading and cells, which is applied in the field of biomedicine, can solve the problems of cumbersome screening of hybridoma cells and difficulties in seed preservation, and achieve the effects of reducing the cost of early tumor intervention, reducing the size of the cell culture system, and reducing the amount of use

Pending Publication Date: 2021-10-12
浙江圣希澳医学科技有限公司
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Problems solved by technology

The hybridoma cell method is gradually eliminated by researchers because the screening of hybridoma cells is cumbersome and difficult to preserve.

Method used

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  • BsAb in-vitro loaded T cell

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Embodiment Construction

[0059] BsAb loading T cells in vitro is characterized in that: the specific method includes:

[0060] S1: BsAb structure design and optimization; design single-chain bispecific antibodies with different configurations, and optimize the sequence and structure;

[0061] S2: Preparation of BsAb; the preparation methods of the BsAb include hybrid hybridoma technology, chemical cross-linking method and genetic engineering method; due to the low yield of hybrid hybridoma technology, time-consuming and laborious, difficult to screen, chemical cross-linking method is easy to cause Protein denaturation cannot activate the antibody, so we plan to use genetic engineering. In order to ensure the function of BsAb, we intend to use eukaryotic expression system for expression.

[0062] S3: Stability analysis of BsAb; take 10ug of the double antibody and place it at 4°C for 7 days, then detect the size of the double antibody fragment by non-reducing SDS-PAGE gel;

[0063] S4: affinity analy...

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Abstract

The invention belongs to the field of biological medicine, belongs to the technical field of cell therapy, and particularly relates to a BsAb in-vitro loaded T cell. The method the following specific steps of structural design and optimization of BsAb; preparation of BsAb; carrying out stability analysis on the BsAb; carrying out affinity analysis on the BsAb; culturing immune cells; loading T cells on the BsAb in vitro; killing in-vitro tumor cells by the BsAb-loaded T cells; secreting cell factors; expression of a proto-oncogene and a cancer suppressor gene; killing tumor cells in vivo by the BsAb-loaded T cells; adopting a TaqMan chip for screening the change of immune related genes and tumor markers in tumor tissues; and detecting the content of CD3 + T cells in the tumor tissue in an immunohistochemical manner. According to the BsAb in-vitro loaded T cells, on the basis of BsAb preparation through chemical cross-linking in the early stage, specific BsAb is planned to be constructed and synthesized, the affinity of BsAb to tumor cell related antigens is increased, an efficient and simple method for loading immune cells in vitro through BsAb is established, and meanwhile an immune cell in-vitro amplification method which is easy and convenient to operate, safe, free of pollution and high in repeatability is established.

Description

technical field [0001] The invention belongs to the field of biomedicine and the field of cell therapy technology, in particular to BsAb loading T cells in vitro. Background technique [0002] In view of the previous establishment of BsAb preparation technology based on chemical cross-linking, it is planned to develop a series of BsAb preparation based on genetic engineering method, and an ultra-early tumor treatment technology that is loaded with autologous immune cells in vitro. This technology has the characteristics of safety, reliability, low toxic and side effects, and strong specificity. It can effectively remove small lesions and prevent tumor recurrence, and provides an important intervention method for ultra-early cancer treatment. [0003] As the latest tumor immunotherapy technology, autologous immune cell therapy has become a new tumor treatment model. Although it has developed rapidly in recent years, the overall effect is still not good. The reason is mainly ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783C07K16/30C12Q1/6886G01N33/574
CPCC12N5/0636C07K16/30C12Q1/6886G01N33/574C12Q2600/158
Inventor 徐健李陶张扬
Owner 浙江圣希澳医学科技有限公司
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