We have been handling animal cell cultures for a long time. Recently, we are facing a problem which we never faced earlier. The problem is that the cell morphology looks pretty good, media is also fine but there are some small particles having slow motion but are not moving away from their place. To remove them, we washed the cells with PBS and changed the medium but next day the particles were back. We took every precaution such as fumigating the lab on proper time, autoclave each item before use etc. There was no contamination in the medium. Incubated flask having only medium and FBS also shown to have such particles. I have attached the video here. Please have a look and give some suggestion to avoid them. Thank you.
I can't see the video but if these objects move I think they could be bacteria or fungi or molds. I suggest you to try to filter the complete medium (medium and FBS) through a 0.22 um filters and verify if the objects will be still present. Maybe the FBS could be contaminated. Try with the filters.
This indeed sounds like you have an infection. My suggestion is that you just prepare new sterile medium, and start a fresh batch of animal cells.
Hi Safiulla, also I can't see the video but from our experience in lab we see like you some particles move slowly in media, this is really an infection and should exclude it and try to prepare another and before use make incubation if these flicks appear try to change your FBS or see your filter in filteration system.
regards
I had a similar experience before. The idea that worked for me was changing serum batch. Get serum samples from couple of companies like GIBCO or Hyclone. Test them and stick with the same batch (including lot no.) for the entire project. Also, if you are preparing your media in-house, then I would suggest using Milli-Q water, which has been autoclaved twice. Another suggestion is to test for mycoplasma contamination. Best of luck.
I also can't see the video, but in my experience what you describe in contamination. Even if you wash with PBS you won't be able to remove it. I agree with previous suggestions, prepare new media, filter it and prepare a new and fresh cell culture. There is nothing else you can do. Good luck!
I thank you all for your valuable suggestions. As you couldn't able to see the video, I have attached the picture here please have a look. It is K562 cell line.
Hello
If the observed particles are moving it means that your cells have been contaminated probably by buffer or trypsin solution. As suggestions
1- Put a small amount of medium with buffer and then incubate to see if the color of the medium has changed, if yes change the buffer
2- You can also do the same with trypsin solution
It sounds like there is an infection in your cell culture. To test this, make new complete media without antibiotics and grow the cells in it for one week. Infections may be hold back by antibiotics, but is still there and will boom when the antibiotics are taken away. If there is an infection the best way of having a healthy cell culture would be to start with fresh media, fbs, pbs etc. and a new batch of your cell line (e.g. if you have frozen aliqotes of it). Good luck!
hello,
it seems that your culture has been contaminated. sometimes it takes a few days to see a contamination grow so as to change the medium ph-colour, or to make it blur. I would suggest to start with every medium or flask etc fresh and new. if your trypsin stock or your antibiotics of FBS stock are aliquoted you could filter them before use in sterile minifilters. if you flame-burn the flask or the medium bottle make sure that no chock residues drods in the bottle. good luck
I cannot see the movie, but anything moving in your culture that is not a cell can be an infection...
Hi, I have faced a similar problem with my cells. I think that is mycoplasma contamination. The only solution for it is to trow away that flasks and start a fresh however you should not use antibiotics while reviving the frozen stocks. I stongly believe that almost all the cells that we grow in laboratory are having this endoparasite, however it breaks out only when the cells are put under stress, especially if we use antibiotics in the media while reviving the frozen stocks.
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