What should I pay attention to when entering the cell?

1. Take and put things from the refrigerator quickly, and check whether the door is closed when closing the door. Store reagents by category.

2. The mother liquor of the culture solution is public, prepared by the students on duty and stored at -20℃ for a long time. When in use, serum is added in proportion to make culture medium working solution, and the used mother liquor is opened and stored at 4℃, so as to avoid pollution. If it is found that the culture liquid stored at 4℃ is insufficient, it will be thawed from -20℃ in time. If there is no reserve at -20℃, please inform the students on duty in time.

3. The subpackaged serum is stored at -20℃ for a long time and pre-thawed at 4℃ before use. Those who take serum from -20℃ need to register and sign on the serum tube. If they use other people's thawed serum, they need permission to avoid delaying other people's experiments. Thaw as needed to avoid repeated freezing and thawing.

4. In principle, all thawed serum should be made into serum-containing culture medium for later use. If there is any surplus, it should be temporarily stored at 4℃ and used as soon as possible. Try not to use other people's serum to avoid cross-contamination.

Sixth, the duties of the students on duty

1. The weekly student duty starts on Saturday and ends next Friday.

2. What should be done at the beginning of duty week: Prepare sterilized gun heads, centrifuge tubes and dried glassware on or before Saturday and Sunday for use next week. Prepare 75% alcohol disinfection, make alcohol cotton balls, and add 95% alcohol to the cell room of alcohol lamp.

Note: Before sterilizing the new centrifuge tube, rinse it with deionized water for 2-3 times, then rinse it with double distilled water 1 time, dry it, wrap it in kraft paper and sterilize it.

Note: 75% alcohol can be prepared by adding 750 ml of 95% alcohol to 950 ml of deionized water.

3. What needs to be done every day during the duty week: turn on the big ultraviolet light to disinfect the cell15-30 min; Lift the plastic bucket full of soaked articles out of the battery room for cleaning, and put the empty bucket back into the battery room; Replace garbage bags; Check whether there is enough water in the incubator; Check the storage of sterilized items in order to replenish them in time; Check the storage of common reagents such as mother liquor, serum and alcohol in culture solution.

4. What needs to be done before the end of duty week: clean the cell on Friday or the day before and after. Including mopping the floor, cleaning desks and cabinets, cleaning drawers, cleaning ultra-clean desks, refrigerators, table instruments, adding or changing water to water bath pots, washing laboratory clothes and special slippers for cell rooms, and sterilizing cell rooms with ultraviolet rays. Remember to replace the incubator with MilliQ water to ensure the water quality is clean!

Note: If the students on duty filled the water tank in the incubator in time last week, according to experience, the incubator will not dry next week, but the students on duty should always check and find out the abnormal situation in time.

5. Clean the incubator every two weeks: firstly, take out the items in the incubator and temporarily store them in the ultra-clean table. Sensitive cells can be temporarily stored in an incubator in the cell room 404; Take out the stainless steel plates in the incubator, including four horizontal plates and two vertical frames, and clean the steel plates and the inner wall of the incubator with alcohol cotton balls; When you open the incubator to clean the inner wall, you should act quickly to avoid a large amount of unclean air from entering the door for a long time and shorten the life of the filter; Irradiate the inside of the incubator with ultraviolet rays for 15 minutes, and wipe the portable ultraviolet lamp with alcohol cotton before putting it into the incubator.

6. Preparation of mother liquor of culture solution: There are two kinds of culture solution: RPMI 1640 and DMEM, and the formula is shown in the liquid preparation room of 404 Palace. Prepare the culture medium, dry bake the measuring cylinder, beaker, 250ml serum bottle, moist heat sterilization filter and serum bottle cap in advance, and soak the domestic filter membrane 15min or overnight with MilliQ water.