Probe domestic double-head probe 038-BB-5.7L The shape of the two ends of the needle is outside the pointed needle
(1) Depletion or deficiency or destruction of essential components such as glutamine or growth factors in some culture cells due to replacement of different culture fluids or serum (2). (3) There is a small amount of bacterial or fungal contamination in the culture (4) Improper preservation of the reagents (5) The initial concentration of the inoculated cells is too low (6) The cells are aging (7) Mycoplasma contamination Shanghai Jinma Biotechnology gives recommendations based on testing experience Method (1) Compare the composition of the new culture medium with the original culture medium, and compare the new serum with the old serum to support the cell growth experiment. Let the cells gradually adapt to the new medium. (2) Replace with freshly prepared culture medium, or add glutamine and growth factors. (3) Incubate with an antibiotic-free culture solution, and discard the culture if contamination is found. Or use antibiotics to remove bacteria. (4) Serum should be stored at -5 ° C to -20 ° C. The culture solution should be stored at 2-8 ° C in the dark. The serum-containing complete medium was stored at 2-8 ° C and used up within 2 weeks. (5) Increase the initial concentration of the inoculated cells. (6) Switch to new seed cells. (7) The culture was isolated and tested for mycoplasma. Clean the stand and incubator. If the mycoplasma is found to be contaminated, discard the culture.
Cell considerations:
1. Before the experiment, the sterile room and the aseptic table (laminarflow) were sterilized by UV lamp for 30-60 minutes, the aseptic table was wiped with 70% ethanol, and the aseptic table fan was turned on for 10 minutes. Only begin the experimental operation. Only one cell line was processed per operation, and the medium was not shared even if the medium was the same to avoid error confounding or intercellular contamination. After the experiment was completed, the test articles were taken out of the workbench and the aseptic processing table was wiped with 70% ethanol. The operation interval should be such that the aseptic table is operated for more than 10 minutes before the next cell line is operated. 2, aseptic operation work area should be kept clean and spacious, necessary items, such as test tube rack, straw suction device or straw box can be temporarily placed, other laboratory supplies should be removed after use, in order to facilitate the circulation of air. The test article was wiped with 70% ethanol before being brought into the aseptic workstation. The experimental procedure should be in the central sterile area of ​​the lifting surface and not in the non-sterile area of ​​the edge. 3. Carefully use sterile laboratory items to avoid contamination. Do not touch the tip of the pipette or the mouth of the container, and do not operate the experiment directly above the open container. After the container is opened, hold the bottle cap with your hand and hold the bottle body, and use it at an angle of about 45°. Try not to place the cap on the table with the cap cover facing up. 4, the staff should pay attention to their own safety, must wear the lab coat and gloves before the experiment. Special care should be taken for cell lines from human or viral infections and an appropriate level of aseptic table (at least Class II) should be selected. During the operation, avoid the generation of aerosol, beware of toxic drugs such as DMSO and TPA, and avoid damage from sharp needles. 5. Regularly check the following items: CO2 pressure of 5.1 CO2 cylinder 5.2 CO2 incubator CO2 concentration, temperature, and water tray for contamination (water for water tray for sterile water, weekly replacement). 5.3. Airflow pressure in the aseptic operating station, regular replacement of UV lamp and HEPA filter membrane, pre-filter (300 hours / pre-filter, 3000 hours / HEPA). 6. The sink can be added with disinfectant (Zephrin1: 750), and the water in the sink should be replaced regularly . After the powder medium is prepared (with serum), it should not be more than 1 month at 4 degrees, such as storage time at -20 degrees. It can be longer, but it should not be more than 3-4 months. It may not be too high for immortalized cell lines, the cells are delicate, and the placement time should not be too long. 7. When the ultraviolet irradiation station is turned on, the medium, the enzyme, and the DHANKS solution are allowed to go outside to allow it to naturally heat up. After 40 minutes, the temperature also rose. Many people put it in a 37 degree water bath to heat it. Be sure to pay attention to the hygiene of the water bath. Some of them are not cleaned all the year round. The inside is very dirty, and it is easy to adsorb a large amount of bacteria on the outer bottle. Therefore, you must change the water frequently when you use it. After taking it out of the water bath, it is best to find a towel to dry the water.
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