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Aspergillus in cell culture room - HELP!!! how hardcore to decontami


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#1 k8e

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Posted 18 July 2010 - 07:19 PM

Hi Everyone,

I am in great need of assistance regarding some contamination in our cell culture room. I am in the final months of my research of my PhD and now am having some huge problems
Recently, I have started sharing the tissue culture room with some newbies (From a different lab) to cell culture (I myself have been doing cell culture for a long time) and it appears as though they have regular contamination issues.. I use a separate incubator and a closed flask system and wasn't overly concerned until the last few weeks when my cells have started to look rather unhappy.
I think they are contaminated with Asp but due to the separate incubators I wasn't sure where the contamination came from.. however when I was in the cell lab doing work last week, 2 of the girls found a plate in their incubator which was entirely taken over by aspergillus, they took it to the sink to remove the lid and pour bleach over the plate.. ;) Until I obviously stopped them from doing this and pointed out it was a cell culture lab, any contaminations should be taken away immediately and thrown out and also advised them to throw out their cell lines and begin again (however apparently they dilute their contaminations out.. hmmm interesting right...)

So it appears this is the way they have been dealing with their contamination issues and why they continue to occurs.. so I am assuming that the entire cell culture room is heaving with aspergillus spores.. That they've probably got into my incubator, the fridge etc.. :(

I have been performing cell culture for quite a few years and never had this problem before and am unsure how to approach the cleaning of aspergillus as I have never seen anything like that (in fact aside from the odd bacterial contamination, odd fungus never really had issues with cell culture).. I will also be having a lengthy conversation with their supervisor and implementing some rather stringent rules to be followed in the cell culture lab...

So How hardcore do I need to go with the cleaning - should I be getting the hoods serviced, and the entire room decontaminated and if so with what reagent?? I intend to decontaminate my incubator with 3% Virkon, autoclave all components etc.. Is that enough to get rid of the evil thing???

Any advice would be so greatly appreciated!!

Katie

#2 bob1

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Posted 19 July 2010 - 01:44 PM

Crikey, those new students (?) need some teaching about cell culture...

It would be a very good idea to get the incubators all cleaned and autoclaved, wipe out the fridges and water baths with virkon or trigene too. Make sure you wipe off virkon as it will rust the stainless steel quite effectively if you are not careful. Also, throw out any media and solutions that have been in contact with the contaminated cultures - this will include medium that has been opened and in the fridge with theirs.

At this point you can do a few cultures and see how it goes, but the hoods may (more than likely will-) be contaminated too, so keep a close eye on all the cultures in the lab and if they show signs of contamination get the hoods cleaned properly - decontaminated, HEPA filters changed etc.

It would also pay to check on the LN2 storage just to make sure that any cultures they may have stored in there are clearly labelled or thrown away if known or suspected to be contaminated.

#3 k8e

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Posted 19 July 2010 - 04:12 PM

Thanks for that.. yeah don't worry all of the media etc has been thrown away.. They are new students and i'm going to talk to their supervisor and get them to go and see the cell culture queen here at the uni.. She'll sort them out!

Obviously if I don't have to get the hood serviced I won't... But at the first signs of anything dodgy and i'll get it professionally decontaminated..
Good idea with the frozen cells, I'll go throw them out too... lucky I like to keep lots of vials from different dates in storage!

Will bleaching all the surfaces in the room, sufice or should I do that with Virkon too??

Cheers,

Katie

#4 HomeBrew

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Posted 19 July 2010 - 05:11 PM

I don't do cell culture, but it seems you have a lot of cleaning work ahead of you... Why don't you get the girls who handled their stuff inappropriately to do much of the cleaning under your supervision? Seems like a good chance to drill in the fact that there are consequences to sloppy procedures...

#5 perneseblue

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Posted 19 July 2010 - 06:20 PM

I don't do cell culture, but it seems you have a lot of cleaning work ahead of you... Why don't you get the girls who handled their stuff inappropriately to do much of the cleaning under your supervision? Seems like a good chance to drill in the fact that there are consequences to sloppy procedures...


I agree with HomeBrew's suggestion. And it would be a good opportunity for these new students to learn how to properly decontaminate a tissue culture room. Hands on experience and all that.
May your PCR products be long, your protocols short and your boss on holiday

#6 bob1

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Posted 20 July 2010 - 03:58 PM

Excellent idea, I think you could probably get away with just cleaning the benches and shelves, I wouldn't worry about the walls too much, unless you hang stuff used in the hood from them.

#7 k8e

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Posted 20 July 2010 - 04:37 PM

Thanks everyone!! Don't worry i'll make them clean it.. Also thinking of moving to another cell culture room as I can't risk their poor technique contaminating my work again!

Cheers,

Katie




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