cell irradiation culture
["Collaborating Experts | Qingqiang Lei, M.S.", "Immunology Sichuan University"], ["Reviewed by | Dr. Lei Wang", "Biochemistry Molecular Biology Zhejiang University"]
Summary
Cell irradiation refers to the process of irradiating cells with ionizing rays such as X-rays, B and y-rays.
Principle
Irradiation creates charged particles or ions that alter the DNA so that lymphocytes lose their ability to proliferate, but still have the ability to stimulate.
Appliance
Cell irradiation will prolong the cell cycle and induce chromosomal aberrations, thereby causing damage to tissues and cells For some cells that proliferate too fast, irradiation can slow down their proliferation.
Operation method
Cell irradiation culture
Principle
Irradiation creates charged particles or ions that alter the DNA so that lymphocytes lose their ability to proliferate, but still have the ability to stimulate.
Materials and Instruments
Cell Culture Dish, Pipette Gun, Pipettes
Fetal bovine serum (FBS), 1640 medium, irradiator
Move
1. Cell culture
1) Take out the seed-preserving cells from the liquid nitrogen tank, quickly put them into a 37 ℃ water bath, and shake them gently to accelerate cell lysis.
2) Mix the cell suspension with 5 mL of prepared medium in a 15 mL centrifuge tube and centrifuge at 1200 rpm for 3 mins.
3) Discard the supernatant, resuspend the cell pellet with 10 mL of medium, add it to a 10 cm cell culture dish, and put it into a 37 ℃ constant temperature incubator for culture.
2. When the cells are in good condition, irradiate the cells with different doses according to the requirements of the experiment.
There are two air conditioners in the room of the irradiator, you need to turn on the two air conditioners before use, the temperature is already set, you only need to turn on the remote control; next to the irradiator there is a dehumidifier, you need to check the dehumidifier before use, if it shows that the water is full of water need to be poured out (irradiator room opposite the room there is a sink can be used to pour the water).
(1) Insert the key, wait for the instrument to power on, the screen displays as Figure 1 interface, click WARM UP, Figure 2 interface, click STRART, the machine begins to warm up, it takes about 20 min;
(2) After warming up, the interface of Fig. 3 will appear, click on MAIN, the interface of Fig. 4 will appear, click on MANUAL, the interface of Fig. 5 will appear, the parameter of voltage and current are fixed value, no need to change, only need to change the time needed for the experiment, the unit of time is second, click on START to start irradiation.
3. After irradiation, continue to put the cells into the 37 ℃ constant temperature incubator for cultivation.
Caveat
1. Inside the irradiator, there is a slot at the top for copper plates, there are three copper plates in total, which can be selected according to the irradiation needs. 2;
2. when irradiating, make sure that the door is closed and that the door handle is in a vertical position, as shown in Figure 6, and the same applies to preheating; if the door is not closed for preheating or irradiation, the machine will display a program error;
3. After the last person has used the machine for the day, wait for the machine to cool down and turn it off, then return the key to the machine itself. The key is normally placed in the third drawer of the row of drawers on the right-hand side of the entrance to room 234, near the cabinet. 4;
4. There are 5 levels in the irradiator. Generally, mice are irradiated at level 1, with three copper plates and no reflector (that is, the heavy black box), and the irradiation time is calculated according to the irradiation dose; and cells are irradiated at level 3, with one copper plate or no copper plate, and the irradiation time is calculated according to the irradiation dose.
Common Problems
1. A large number of cells die after irradiation, indicating that the radiation is too strong;
2. set up a good control group with a gradient of radiation intensity.
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