Equipment: ① Syringe Reagents: ① IMDM; ② 1x PBS; ③ Acidified antibiotic water.
Move
The basic process of bone marrow cell transplantation experiments can be divided into the following steps:
(i) Preparation of Bone Marrow Transplant Recipient Mice
A Recipient mice are fed with sterilized water acidified with antibiotics for 1 week prior to radiation exposure.
B Recipient mice should continue to be fed acidified sterilized water with antibiotics for 2 weeks after radiation exposure before switching to acidified drinking water without antibiotics.
C For C57BL6/J mice, the dose of radiation is 10 Gy of gamma radiation, and the recipient mice undergo tail vein bone marrow transplantation within 4 hours.
(ii) Preparation of Donor Bone Marrow Cells for Transplantation
A Donor mouse femur is harvested and preserved in RPM11640 (2% FBS, 10 U/mL heparin + penicillin + streptomycin).
C The bone marrow cavity of the femur was rinsed with a syringe and the bone marrow cells were filtered using a sterilized 40 μm cell filter.
C The cells were washed with serum-free RPMI1640 and centrifuged at 4 ℃ for 5 minutes at 2000 r/mi.
D Cells were resuspended in 25 mL of serum-free RPMI and counted.
E Serum-free RPMI1640 was used to adjust the cell concentration to 1x10 cells/mL.
F Bone marrow cells were injected into the tail vein of recipient mice. 0.5 mL of cell solution was injected into each mouse, i.e. 5x106 cells.
(iii) Acidified Antibiotic Water Preparation
A Adjust the pH of water to 2.6 with HCl and sterilize.
B Add 10 mL of neomycin (10 mg/mL) and 400 μL of polymyxin B sulfate (25 mg/mL) per liter of acidified water, filter and sterilize.
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