Human monoclonal antibody of human endophloeodal growth factor receptor 2 protein and its preparing process
An endothelial growth factor and monoclonal antibody technology, applied in the field of biomedicine, can solve problems such as the inability to fundamentally solve the HAMA problem, and achieve the effects of improved effect, long half-life and high yield
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Embodiment 1
[0039] Example 1: Preparation of fully human HER2 / neu monoclonal antibody by obtaining sensitized B cells from peripheral blood
[0040] Step ①, first determine the target volunteers, select 100 volunteers, all of whom are breast cancer patients, take 5ml of venous blood each, separate the serum, store at -20°C, and screen through the commercial HER2 ELISA KIT, the anti-HER2 antibody is positive and the titer Those whose ratio is greater than 1:64 are the target volunteers; then take 50ml of peripheral blood from the target volunteers, add lymphocyte stratification solution, centrifuge at 200×g, take the interface B cell layer, and take 100ul B cells for immune bead test to determine whether the B cells are sensitization. The sensitized B cells were combined and washed 3 times with RPMI1640. Step ②, use RPMI1640 to regulate B cells to 5×10 6 / ml, human myeloma cells adjusted to 2×10 for Karpas707H RPMI 6 / ml. Mix Karpas 707H with 1ml of sensitized human B cells, add 2ml of...
Embodiment 2
[0041] Example 2: Preparation of fully human HER2 / neu monoclonal antibody by obtaining sensitized B cells through in vitro immunization
[0042] Step ①, purchase 400ml of freshly collected venous blood anticoagulated with sodium citrate from the blood bank, separate the plasma, confirm that it is negative for anti-HER2 antibody by commercial HER2 ELISA KIT, add lymphocyte stratification solution, centrifuge at 200×g, and take the interface B cell layer. Take 100ul B cells for immunobead test to determine that the B cells are not sensitized. Unsensitized human peripheral blood B cells were cultured in conditioned medium for 24 hours, then transferred to complete medium for 5 days. The complete medium contained HER2 / neu extracellular region 2ng / ml, AB type human serum 1%, IL -2 100U / ml, IL-4 8ng / ml, IL-6 16ng / ml, macrophage 1×10 5 / ml, 28% of the B cells were sensitized as determined by the immunobead test, and the sensitized B cells were washed 3 times with RPMI1640; in step ...
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