bacterial or fungal contamination - have no clue !!!! (Dec/14/2005 )

I had received the HeLa cells last week and since then i was growing in my lab and had subcultured 3 times.

Since yesterday, i am noticing some rounded stuff , black (very dark) in color.

Is it some contamination ?????????????? or...

those r dead cells.

In case, its a contaminastion, how do i confirm it and what do i do to keep the incubator and other stuff clean. Any special precaution


Plesae advise.
thanks

Hi

First question, how big is the suspect stuff? if it is tiny (400X) and moving rapidly it will be bacteria, if it is large and fuzzy/hairy it is fungal.

Black bits are often cell debris, especially from senescent cells (from my experience).

You can test by taking some of the medium and spreading it on an agar plate and seeing if it grows.

to keep the incubator clean, regularly remove and decontaminate the water pan and replace water, this should be done every week. About once a month remove and autoclave the shelved and any other removable bits. Decontaminate the inside of the incubator by wiping with virkon to kill spores, then with water to remove the virkon, and finally with ethanol. Spray shelves and other removable bits with ethanol and replace.

Good luck
Bob

[quote name='bob1' date='Dec 15 2005, 10:58 PM' post='34681']
Hi.

Well bacteria are going to be itty bitty in comparison to your cells, like the other reader mentioned. They usually appear to be small polk-a-dots that are EVERYWHERE - the stuff spreads pretty effectively. The biggest indicator of bacterial contamination is cloudy media - the media will be very turbid. Normally when your cells are attached to the flask / well, it should be crystal clear (this is of course for adherent cells, like HeLa's.) Fungus however will be long and filamentous looking - like branches on a tree. The media will not be cloudy and you may find some of your cells attached to branches of the filaments.

I have also found that black bits tend to be cellular debris.


Best way to test - plate it. But I wouldn't do it in the same vicinity as your cells. I'd go to another lab altogether - if it is contamination, you want to take every precaution to keep it from spreading.

best way to avoid contamination is asceptic technique and REGULARLY CLEANING lab equipment!!!!! H2O bath