I am working with iPSC cell culture and finding contamination after two or three days. Media color changes yellowish green and turbid. I was not able to see any cells after the contamination even after 70-80%confluency. I am attaching pictures of the plate.
Looks like a bacterium. The best way to solve that is to fix your technique. It's getting in there somehow. You just need to spot how. As a second choice, you could try different antibiotics. Half strength pen/strep can be used prophylactically. Harsher compounds like clarithromycin, erythromycin, ciprofloxacin, are going to impair your iPSC and should only be used in emergencies.
Hi!
This look like bacterial contamination. you can place the plate under the the microscope and be able to see bacteria floating around. Just to be 100% sure i would use this well to perform mycoplasma check and then trash the plate since at 37 whatever is there might spread and contaminate other plate.
Hello Puja Soni
I too feel it is bacterial contamination. You may confirm by streaking a few microlitres of the culture on LB agar plate and leaving the plate at 37 degrees C overnight. If you find bacterial colonies on the plate the next day, it means presence of bacterial contamination.
I would not recommend treating the contaminated culture with antibiotics. In the process of getting rid of bacterial contamination, you would be doing more harm than good to the cells. It is better to discard the plate immediately as you would be putting other cultures in the incubator at risk.
The best way out would be to strictly maintain a sterile environment while working with cells.
Best.
Looks like bacterial contamination. Fungal contaminations take more than 3 days to show. If the contamination is confined to only few wells, then the trouble may be in your handling process. If not, then probably something that all wells share in common (maybe the media, pipette tips, PBS, etc) may be contaminated. In such case, autoclaving all solutions and re-sterilisation of all facilities is suggested.
It does look like bacterial contamination. Even after proper aspetic techniques if the contamination persists, look at sterilizing the incubator, changing the HEPA filter
Hi,
I actually dealt with the same problem as you in the last two months. You can read about it here: https://www.researchgate.net/post/How_to_deal_with_bacterial_contamination_in_iPSC_cultures/1.
After two or three passages, bacteria would contaminate my iPSC cultures, causing me a great deal of inconvenience.
Despite sterilising our L2 room and scrutinising our medium and reagents, the issue persisted.
Consequently, I resorted to antibiotics, experimenting with both the mix of penicillin/streptomycin and gentamycin as well as pure antibiotic with normocure. Fortunately, both approaches yielded positive results, and the situation is now under control.
Nevertheless, I remain uncertain whether the contamination is truly at fault. I am striving to maintain the cultures at the lowest possible spontaneous differentiation percentage. This practice has evidently enhanced the cells' overall wellbeing and prevented sudden contamination by exercising caution.
I cannot definitively state whether there is a correlation between the pluripotency state of the iPSC and the occurrence of contamination. However, what I can say is that the healthier the cells are, the lower the rate of contamination.
I hope this could help you.
Good luck!
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