Macrophage culture method

Macrophage is located within tissue leukocytes, monocytes derived cells, and monocytes from the bone marrow and the precursor cells. Macrophages and monocytes are phagocytic cells, non-specific defense in vertebrates (innate) and specific defense (immune). Their main function is to fix the cells in the form of free cells or cell debris and the phagocytosis of pathogens (i.e. phagocytosis and digestion), and activated lymphocytes or other immune cells, making it react to pathogens.
Culture methods
Macrophage cell line also has unlimited, mostly from the mouse, such as P338D1, S774A.1, RAW309Cr.l, etc., were vicious, macrophages were cultured in cell morphology and phagocytic function, easy passage and sidewall separation, but difficult to construct strains. Cultured macrophages in various ways and from various sources available to get the cells to mice derived method is most useful, and its method is as follows:
1, three days before the experiment, each mouse was intraperitoneally injected to thiolacetic acid sterile broth 1ml (Do not inject the intestines).
2, hoping to kill animals, portable mouse tail will all be immersed in 70% ethanol for 3 to 5 seconds.
3, set the animal on the autopsy table, with a needle fixed limbs, hands holding tweezers pull to the side torn skin, exposing the peritoneum, but not to hurt the peritoneal wall.
4, and then 70% alcohol scrub peritoneal wall, the use of a syringe was injected into the abdominal cavity suction 10mlEagle while rubbing pressure from both sides of the peritoneal wall with your fingers, so that adequate flow of fluid in the abdominal cavity.
5, gently stir the abdominal wall with a needle, the animal body slightly to one side, so that the fluid collection in the abdominal cavity under the needle into the needle tube drawing.
6, carefully pull out the needle, inject the liquid centrifuge tube, 250g4 ℃ for 10 min, the supernatant, add 10mlEagle medium.
7, cells were counted per mouse can produce 20 ~ 30 × 105 cells, of which 90% macrophages.
8, to obtain tie with 3 × 105 cells / square centimeter, to be inoculated with 2.5 × 106/ml. 9, the cell culture was purified to remove other white blood cells, a few hours after inoculation, culture solution, rinsed with Eagle 1 to 2 times, the addition of a new culture medium Eagle set 37 ℃ CO2 culture incubator.

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