Lentiviral vector for expressing CD19 and CD20 antibody gene CARs (chimeric antigen receptors) and preparation method of lentiviral vector

A chimeric antigen receptor and lentiviral vector technology, applied in the field of medical virology, can solve the problems of time-consuming, difficult transfection, low reproducibility, etc., and achieve the effect of wide application range, high infection efficiency and high reproducibility

Inactive Publication Date: 2017-05-31
刘晓明
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Problems solved by technology

[0006] At present, the following problems are encountered in the production of recombinant lentiviruses expressing CD19 and CD20 combined antigen receptors: 1) long time-consuming; 2) low repeatability; 3) difficult or even impossible to transfect; 4) difficult to achieve target gene expression Efficient expression, some protein-related functional experiments are difficult to carry out

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  • Lentiviral vector for expressing CD19 and CD20 antibody gene CARs (chimeric antigen receptors) and preparation method of lentiviral vector
  • Lentiviral vector for expressing CD19 and CD20 antibody gene CARs (chimeric antigen receptors) and preparation method of lentiviral vector

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Embodiment Construction

[0044] The technical solution of the present invention will be further described below, but it is not limited thereto. Any modification or equivalent replacement of the technical solution of the present invention without departing from the spirit and scope of the technical solution of the present invention should be covered by the protection scope of the present invention middle.

[0045] A method for preparing a lentiviral vector expressing CD19 and CD20 antibody gene chimeric antigen receptors, comprising the following steps:

[0046] S1: Preparation of 293T cells

[0047] a. recovery of 293T cells from the initial cell bank;

[0048] b. Suspend the revived 293T cells in 10% FBS DMEM medium (containing 4.5g / L glucose, 1% sodium pyruvate, 1% glutamine), press 2X10 4 / cm 2 The inoculum amount was inoculated in a T75 culture flask, placed at 37°C, 5% CO 2 incubator culture;

[0049]c. Passage and expand the 293T cells every 3 to 4 days. The passage method is as follows:

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Abstract

The invention discloses a lentiviral vector for expressing CD19 and CD20 antibody gene CARs (chimeric antigen receptors) and a preparation method of the lentiviral vector. The lentiviral vector for expressing the CD19 and CD20 antibody gene CARs is formed through cotransfection of 293 T cell by a shuttle expression vector and an auxiliary packaging plasmid, recombination harvest and concentration, wherein the shuttle expression vector contains CD19 and CD20 antibody gene CAR genes. The preparation method of the lentiviral vector expressing the CD19 and CD20 antibody gene CARs is simple, feasible, short in consumed time and high in repeatability; a produced lentivirus has high infection efficiency, can express a target gene efficiently, causes little immune response of a body and is quite wide in application range.

Description

[0001] Technical field: [0002] The invention relates to the technical field of medical virology, in particular to a lentiviral vector expressing CD19 and CD20 antibody gene chimeric antigen receptors and a preparation method thereof. [0003] Background technique: [0004] Lymphatic T cells are the main components of cellular immunity, and using genetic engineering to make T cells express chimeric antigen receptors can transform T cells into effector cells that can specifically kill tumor cells, which is a promising strategy for tumor immunotherapy . The key to the application of chimeric antigen receptors is to determine a tumor-associated antigen, which has overexpression or high expression on the surface of tumor cells, but no or low expression in normal tissues. CD19 is a surface protein expressed on B lymphocytes and follicular dendritic cells, belonging to the immunoglobulin (Ig) superfamily, located on the short arm of chromosome 16 (16p11.2), encoding a 556-amino aci...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/867C12N7/01
Inventor 朱分禄刘晓明
Owner 刘晓明
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