CTLA4lg and derivatives thereof used for resisting bacteria
A derivative, anti-bacterial technology, applied in the direction of anti-bacterial drugs, resistance to vector-borne diseases, medical preparations containing active ingredients, etc.
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Embodiment 1
[0058] Example 1 Study on the regulatory effect of CTLA4Ig on the toxicity of human peripheral blood mononuclear cells (PBMC)
[0059] 1. Reagents and materials:
[0060] Human PBMC: from the Blood Transfusion Department of Chongqing Southwest Hospital, manual separation of concentrated white blood cells;
[0061] CTLA4Ig, Recombinant Human CTLA-4 / Fc Chimera, Catalog Number: 325-CT, commercially purchased from the American R&D system company;
[0062] Human Lymphocyte Separation Solution 200ml, Catalog Number: LTS1077, purchased from Tianjin Haoyang Biological Products Co., Ltd. (TBD);
[0063] CFSE (carboxyfluorescein diacetate succinimidyl ester), Catalog Number: ALX-610-030, commercially available from Alexis biochemicals;
[0064] Propidium Iodide / propidium iodide, Catalog Number: 537059, commercially available from Calbiochem, San Diego, CA;
[0065] K562 (human chronic myelogenous leukemia cell line): purchased from the Shanghai Cell Bank of the Chinese Academy of Sci...
Embodiment 2
[0082] Example 2 CTLA4Ig effectively promotes the ability of human NK cells to kill sensitive target cells (≥1 μg / ml)
[0083] 1. Reagents and materials:
[0084] Human NK cell isolation kit (NK Cell Isolation Kit human), Catalog Number: 130-092-657, commercially available from miltenyi biotec (Germany).
[0085] LD columns, Catalog Number: 130-042-901, miltenyi biotec
[0086] All the other materials are the same as in Example 1.
[0087] 2. Experimental method:
[0088] The separation of human PBMC is the same as in Example 1;
[0089] Human NK cells were isolated and operated strictly according to the kit instructions;
[0090] Cytotoxicity detection method:
[0091] The method of CFSE labeling human NK cell sensitive target cell K562 is the same as that in Example 1.
[0092] Co-culture of effector cells and target cells: 10 5 Personal NK cells +2×10 3 A CFSE-labeled K562 was added to a 96-well plate and co-cultured for 2.5 hours. K562 without PBMC was used as the...
Embodiment 3
[0101] Example 3 CTLA4Ig effectively promotes the ability of human NK cells to kill sensitive target cells (<1 μg / ml)
[0102] 1. Reagents and materials:
[0103] With embodiment 2.
[0104] 2. Experimental method:
[0105] The separation of human PBMC is the same as in Example 1;
[0106] Human NK cells were isolated and operated strictly according to the kit instructions;
[0107] Cytotoxicity detection method:
[0108] The method of CFSE labeling human NK cell sensitive target cell K562 is the same as that in Example 1.
[0109] Co-culture of effector cells and target cells: 10 5 Personal NK cells +2×10 3 A CFSE-labeled K562 was added to a 96-well plate and co-cultured for 2.5 hours. K562 without PBMC was used as the natural death control of target cells. K562 with IL-2 added was used as a positive control.
[0110] The methods for calculating the killing efficiency of PI-labeled dead cells, flow cytometry detection and NK cells are the same as those in Example 1. ...
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