My MCF 7 cell line in my co2 incubator after 24 hr either subculture or thawing getting turbid and flask media having particles which pellet down. What is the problem kindly suggest.
Hello Arpita Dey
How well-versed are you in cell culture? Are you maintaining proper sterile condition while working in the cell culture laboratory? When you work with cells in tissue culture it is not just about handling cells but you also need to take all the necessary precautions to avoid contamination. Contamination in cell culture is a serious issue and you need to avoid it as far as possible. If your culture gets contaminated, other cultures kept is same incubator are also likely to get contaminated.
So, I would suggest you get your incubator cleaned. Get all the trays and the water tray autoclaved. Using a lint free cloth, clean the chamber interior with soapy water and rinse with water, followed by wiping the surfaces with alcohol (70 %) or an equivalent non-corrosive disinfectant. Change the incubator water (not just refill it, but empty and add fresh, sterile, distilled water) at least every other week. Clean the incubator one to two times per month (depending on the number of users). It is not necessary to autoclave everything. Wipe down the incubator with 70% ethanol, especially the water pan (do not spray ethanol on sensors). Allow to air dry. Do not store anything on top of the incubator and clean the top of the unit every two weeks to remove dust. Wipe down the doors and handles with 70% ethanol. Also, clean all spills immediately.
Also, do a sterility check of your culture media. Growth media should always be tested for sterility by placing it in a 37 degree C CO2 incubator for 72 hours prior to utilization to ensure that the media is contamination-free.
Best.
Either media or cells are potentially contaminated. Do you use antibiotics in media? Cavicide is sometimes better to use than alcohol. Do you keep your waterbath clean? How do you thaw the cells? Growth check of media is definitelly necessary to do. Where did you get the cells from? Were they checked for sterility?
A turbid appearance and the presence of particles in the media of your MCF-7 cell line can be indicative of contamination. Here are a few possible causes and suggestions for troubleshooting:
1. Bacterial or fungal contamination: Turbidity and the presence of particles in the media can occur due to bacterial or fungal contamination. Contaminating microorganisms can multiply in the culture and cause changes in the media appearance. To address this, perform a thorough check of your laboratory practices and ensure proper aseptic technique during cell handling and media preparation. Use sterile techniques, including clean workspaces, proper disinfection of surfaces and equipment, and appropriate handling of reagents and media.
2. Mycoplasma contamination: Mycoplasma is a common type of cell culture contaminant that can cause turbidity and changes in media appearance. Mycoplasma contamination often goes undetected because it cannot be seen under a microscope. To confirm or rule out mycoplasma contamination, perform regular mycoplasma testing using commercially available kits or other validated methods.
3. Autoclave issues: If you autoclave your media or other components, problems with the autoclave cycle or incorrect sterilization can lead to contamination. Make sure you are using appropriate sterilization methods and check the autoclave settings, such as temperature and duration, to ensure effective sterilization.
4. Contaminated reagents or media components: If you are using pre-made media or supplements, there is a possibility that the contamination originated from these sources. Ensure that all reagents, media, and supplements are obtained from reputable suppliers and stored properly to minimize the risk of contamination.
5. Cross-contamination: It's important to confirm the identity and purity of your MCF-7 cell line. Cross-contamination with other cell lines can occur, leading to changes in cell behavior and media appearance. Perform routine authentication of your cell line using methods such as DNA profiling or short tandem repeat (STR) analysis.
In cases of suspected contamination, it is recommended to discard the contaminated cultures and decontaminate your incubator, equipment, and reagents. Start fresh cultures from a known and authenticated source, taking precautions to maintain sterility throughout the process.
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Cell Biology
- Culture Cells
- Cell Line