In my experiment I got contaminated HeLa cell culture plate. I don't know the reason behind it. can anybody please suggest something worthy to resolve such kind of issues?
When it comes to the routine use of antibiotics in cell culture media, I have to disagree completely. Don't use them regularly in your standard cell line cultures. A lot of contaminating bacteria are already resistant against various antibiotics or at least you will promote the formation of resistance. Moreover: You don't need them, if you stick to the basic hygienic rules (see the first comments) and they will give you an feeling of 'false security'.
I would definately clean up the water bath each week by draining the old and using new deionized water, cleaning the surfaces, followed up disinfection with bacteriocidal/antiseptic chemical treatment of the water.
Have you been able to find what type of contamination is it, i.e. bacterial or fungal? It is not unusual to have contamination every-now-and-then, but persistent & widespread contaminations warrant an investigation. Ideally, everyone in the lab would be operating with the use of "aseptic technique". You can find many articles about aseptic technique online. I would suggest the following link for a quick overview;
https://wikisites.mcgill.ca/djgroup/images/9/9d/Contamination_%26_cell_line_mixed_up.pdf
Thank you Kiessling for you nice suggestions. I have followed all measures you just mentioned, by the way I have not done identification yet. I have repeated my experiment and hopefully I won't have any problem in future.
To avoid contamination, I would recommend using 70% ethanol regularly while dealing with culture plates. Spray thoroughly to all the apparatus (such as pipettes, pipetter, reagent bottles).
Make sure that, you open lid of every reagent bottle inside the hood.
Lastly, spray your gloved hands with 70% ethanol as regularly as you can!!!
--Best regards
I should also stress that ideally everyone using the facility is trained by someone competent. As the saying goes, your cell culture facilities are as good as the worst person using it. That means, if someone has a sloppy technique (thus leading to contamination), the consequences can be felt for other people despite the properly trained users obeying strict guidelines.
Thanks Herald for your suggestion, but mycoplasma are typically partially or completely resistant to the antibiotics commonly used in cell cultures such as penicillin
and streptomycin.
Are you having a mycoplasma issue? You will not be able to see this type of contamination, i.e. by microscopy, turbidity and pH changes. It is a very subtle contamination, the easist way to probe for this would be to grow cells for a long time and use DAPI. The good news is that there are treatments available for mycoplasma contamination but they are somewhat expensive and remains useful in a limited setting.
http://weill.cornell.edu/vascularbiology/protocols_and_downloads/pdf/Mycoplasm-Test_HLA.pdf
I think you need to follow all the suggestions mentioned above as well as add a particular amount of wide-spectrum antibiotic into your culture medium that's what I do usually in my work, but be cautious that it depends on your experiment and what you plan to do.
When it comes to the routine use of antibiotics in cell culture media, I have to disagree completely. Don't use them regularly in your standard cell line cultures. A lot of contaminating bacteria are already resistant against various antibiotics or at least you will promote the formation of resistance. Moreover: You don't need them, if you stick to the basic hygienic rules (see the first comments) and they will give you an feeling of 'false security'.
Definately agree with Max, using antibiotics may even mask bad technique and sometimes they rarely accomplish what they were designed to prevent!
Hello, in my experience I just follow aseptic technique, also add 1% antibiotic with media. in addition, before I starting with cell culture spray 70% alcohol and add my consumables in BSC then UV 20min after that u can work without any problems.
good luck
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