What type of contamination is this? - (Aug/14/2016 )

Hi there,

 

I am currently working on two oral carcinoma cell lines. I did all the labworks for my first cell line in my original lab, and everything turned out to be fine. 

 

As for the second cell line, I did the labworks in another cell culture lab (because it was more spacious and lesser ppl were actually using it). But the problem is that I noticed my cells had been contaminated, most likely was due to the media. But I am nt quite sure what kind of contamination it is.

 

I fast thawed my cells, and revived them on 09.08.16, and checked on the the next day (on 10.08.16):

 

Less than 50% of cell viability but I thought it was normal and did not suspect any thing because I stored the cells in -80 instead of nitrogen tank, so I thought it might have affected my cell viability.

After that, I washed away the dead cells:

 

 

 

As you can see, i started to saw the fibre liked thingy besides my cells, which i hoped was just debris from serum, so i filtered my media, and transfered some of the filtered media (without cells) to an empty flask (i'll name it Flask A) as reassurement. It appeared to be clear from any cell or debris bt then funny stuff started to appear on the flask A the next day:

and when i used higher magnification to look at them, they appeared to be something like this:

 

Meanwhile, the second cell lines that i revived on 09.08.16 were growing like this:

and I think the contaminants were consuming cells, because the doubling time for this cell line is less than 20 hours.

 

But I planned to observed for a few more days:

So, on today (15.08.16), all my cells were dead, and fungal-liked thingy were actually growing instead: 

 

where as my flask A (only filtered media with no cells),

 

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when i first transfered the media to the flask, the appeared to be in dark spots:

then after a few days, they looked like the ones i showed in the previous images.

 

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so i throw everything..

Idk what type of contaminant is it and this is my first time encountering it =(

im suspecting the water bath that i used, because since not much ppl are actually using the lab, the water bath was really old and i can see fungi floating on the water when i first planned to use it, i cleaned it few times and added water disinfectant before using it already, but probably it is nt clean enough =(

 

pls help, and do you have any suggestion on encountering such problems? as i m still very new into cell culturing 

Thank you very much in advance..

so i throw everything.. 

The photos of the contaminant (photos 4, 5 and 7) look like you are focused on the outside surface of the flask, photo 7 in particular is a dried drop of medium or serum. 9, 10 and 11 also look like the outside of the plate, the bright spots are dust.

Really? THANK YOU SO MUCH, i will try to readjust and focus using the microscope..

 

But any idea what is the fibre liked thingy? Between my cells.. 

The fibrous things are probably plastic residues from the flask or debris from the FBS.

Thank you very much! 

Your help is greatly appreciated~