Method for quick activation of LAK (Lymphokine activated killer) cells
A technology of lymphokines and killing cells, which is applied in the direction of animal cells, vertebrate cells, cell culture active agents, etc., and can solve problems such as weak lethality, multi-organ dysfunction, and large-scale infusion
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Embodiment 1
[0050] Embodiment 1: the preparation of polypeptide
[0051] 1. Defat wheat germ
[0052] Use defatted wheat germ, or directly purchase defatted wheat germ for subsequent experiments. Degreasing conditions: extractant: petroleum ether (60°C-90°C), water bath temperature: 65°C, solid-liquid ratio: 1:8 (g / mL), degreasing time: 4h.
[0053] The defatted wheat germ is pulverized into 80 meshes to obtain defatted wheat germ powder, which is set aside.
[0054] 2. Hydrolysis and purification of peptides
[0055] The defatted wheat germ powder was treated with papain (enzyme activity of 800,000 U / g) + neutral protease (enzyme activity of 200,000 U / g) at a compound ratio of 3:1 (mass ratio), pH of 6.0, and temperature Enzymolysis under the conditions of 55°C, substrate mass concentration 40g / L, enzyme addition amount 5% (100g substrate plus 5g enzyme), enzymolysis time 8h, after enzymolysis, the temperature was raised to 90°C and kept for 20min for enzymatic inactivation treatment ...
Embodiment 2
[0059] Embodiment 2: LAK cell rapid activation method
[0060] 1. Experimental materials
[0061] RPMI 1640 medium and 10% fetal bovine serum were purchased from Gibco.
[0062] rIL-2 was purchased from Beijing Sihuan Biopharmaceutical Company.
[0063] 2. Experimental method
[0064] 1. Umbilical cord blood collection and mononuclear cell isolation
[0065] The closed collection method was used to collect cord blood from 100 cases of healthy neonates born in full-term normal delivery, and the parents of the prenatal examinations had no family history of genetic diseases. It was collected from the side away from the neonatal side of the umbilical cord stump before delivery. After collection, they were stored in a refrigerator at 4°C, and separated and frozen within 24 hours.
[0066] With reference to the technical handbook in this field (Shen Caring etc., Modern Immunology Experimental Technology, Hubei Science and Technology Press, 1998, 172-175 pages), adopt Ficoll dia...
Embodiment 3
[0080] Embodiment 3: LAK cell rapid activation method
[0081] 1. Experimental materials
[0082] RPMI 1640 medium and 10% fetal bovine serum were purchased from Gibco.
[0083] rIL-2 was purchased from Beijing Sihuan Biopharmaceutical Company.
[0084] 2. Experimental method
[0085] 1. Umbilical cord blood collection and mononuclear cell isolation
[0086] The closed collection method was used to collect cord blood from 100 cases of healthy neonates born in full-term normal delivery, and the parents of the prenatal examinations had no family history of genetic diseases. It was collected from the side away from the neonatal side of the umbilical cord stump before delivery. After collection, they were stored in a refrigerator at 4°C, and separated and frozen within 24 hours.
[0087] With reference to the technical handbook in this field (Shen Caring etc., Modern Immunology Experimental Technology, Hubei Science and Technology Press, 1998, 172-175 pages), adopt Ficoll dia...
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