Preparation of specific tumor killing cell
A cell-killing and species-specific technology, applied in the field of anti-tumor cellular immunotherapy, can solve the problems of low clinical effective rate and low tumor killing rate
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Embodiment 1
[0096] Use the COBE SPECTRA cell component separator, set the parameters and program according to the instruction manual, collect 6000-8000ml of peripheral blood, and collect 120-140ml of mononuclear cells. Divide into 50ml centrifuge tubes, and centrifuge at 1500 rpm for 5 minutes in a Hitachi cell centrifuge. Collect the upper layer of plasma and transfer it to a 50ml centrifuge tube to prepare autologous inactivated serum (method: add 2.5ml of 10% calcium gluconate by pipetting and mixing, bathe in 56°C water for 35 minutes, centrifuge at 2500 rpm for 5 minutes, collect the supernatant), and refrigerate at 4°C spare. Collect the cells, dilute them with 30ml of normal saline, transfer them into a 50ml centrifuge tube containing 15ml of lymphocyte separation medium (Ficoll, 1.077), centrifuge at 2000rpm for 15 minutes in a horizontal rotor centrifuge, and absorb the middle white cell layer (monocytes) with a disposable pipette. ). Take 15ml of mononuclear cells and transfer...
Embodiment 2
[0110] Example 3
[0111] The present invention also includes a combined reagent, which is characterized in that it includes all the reagents needed to prepare specific DC-CIK-CTL cells and preparations thereof, and the names of the reagents are as follows:
Embodiment 3
[0113] 11. Monocyte macrophage colony stimulating factor (GM-CSF);
[0114] 12. Interleukin 4 (IL-4);
[0115] 13. Gentamicin;
[0116] 14. Autologous tumor-associated whole antigen;
[0117] 15. IFN-γ;
[0118] 16. CD3 monoclonal antibody;
[0119] 17. Interleukin-2 (IL-2);
[0120] 18. 0.9% sodium chloride;
[0121] 19. Human albumin;
[0122] 20. Interleukin-2 (IL-2) for injection;
[0123] The doses of the reagents are as follows:
[0124]Doses and final concentrations of DC-CIK-CTL combination reagents are as follows:
[0125] 11. GM-CSF: 50ug / 500ml;
[0126] 12. IL-4: 30ug / 500ml;
[0127] 13. Gentamicin: 20,000 units / 500ml;
[0128] 14. Autologous tumor-associated whole antigen: 50ug / ml;
[0129] 15. IFN-γ: 50ug / 500ml;
[0130] 16. CD3 monoclonal antibody: 25ug / 500ml;
[0131] 17. IL-2: 15ug / 500ml;
[0132] 18. 0.9% sodium chloride: 200ml;
[0133] 19. Human albumin: 2.0g;
[0134] 20. Interleukin-2 (IL-2) for injection: 200,000 U;
[0135] Combination ...
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