Aseptic operation is a key factor that determines the success or failure of cell culture. Even if a well-equipped laboratory is used, if the experimenter is careless and the operation is not standardized, it will also cause contamination. Therefore, all operations must be aseptic as possible, and every work must be orderly and completely reliable.
1. Before the experiment: Make an experiment plan and operating procedures before starting the experiment. The calculation of the relevant data should be done in advance. According to the experimental requirements, prepare various required equipment and items, place them in the operation place (cultivation room, ultra-clean table) after inventory is correct, and then start disinfection. This can avoid increasing the chance of contamination due to incomplete items being taken back and forth after starting the experiment.
2. The working area for aseptic operation should be kept clean and spacious. Necessary items such as test tube racks, straw extractors or straw boxes can be temporarily placed, and other experimental supplies should be removed when they are used up to facilitate the flow of airflow. The experimental supplies were wiped with 70% ethanol before being brought into the sterile operating table. Cell culture experiments should be performed in the central aseptic area of the raised surface, and do not operate in the non-sterile area at the edges. To
3. Take care to use sterile experimental items to avoid contamination. Do not touch the tip of the straw or the mouth of the container, and do not operate the experiment directly above the opened container. After the container is opened, hold the bottle cap with your hands and hold the bottle body, and use it at an angle of about 45°. Try not to put the bottle cap on the table top. To
4. The staff should pay attention to their own safety and perform experiments only after wearing lab coats and gloves. For cell strains derived from humans or virus infections, you should be especially careful and choose an appropriate level of aseptic operating table (at least Class II). During the operation, avoid causing the production of aerosol, be careful with toxic drugs, such as DMSO and TPA, and avoid injury from sharp needles. To
5. Regularly check the following items
(1) CO2 pressure of the CO2 cylinder; (2) CO2 concentration, temperature, and whether the water pan is contaminated (the water in the water pan is sterile water, which is replaced weekly); CO2 incubator is regularly disinfected; (3) aseptic operation Whether the airflow pressure in the station is normal, replace the ultraviolet lamp and HEPA filter membrane, pre-filter (300 hours/pre-filter, 3000 hours/HEPA) regularly.
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