Dear all,
My lab has just recently got a cell line. The cells grow a bit slowly and the medium color is changing quite fast from pink to orange compared to other cell lines. In addition, we observed some strange black debris. However apparently the movement is brownian, unlike bacterial movement.
Have you ever experienced thing like this? Could you help me pinpoint what is this debris and share any suggestion what should I do? Thank you.
Are you using a scrapper at the time of passage? Some times improper handling of scrapper while passage can cause cell fragmentation and these fragments can appear as black spots or excess cell debris.
Khrisna Putera Buddhiman
Hi
There are many possibilities for this to happen and it is sometimes seen in the fields of crops. In some cases, it is seen that change the medium more often with perhaps some optimization of the growth factors and nutrients.
But something that has a very high probability is contamination.
Please, if you are able to, do this test, as well as for everyone else in the globe who is struggling with this issue:
Please add 5 milliliters of serum to the flask (no gas exchange cap or parafilm completely, these bugs grow fast under anaerobic conditions). And incubate at a temperature of 37 degrees (NOT CO2 incubator). After two to three days, examine the cells with a 40x magnification and phase contrast. (do not use 20X magnification or bright field), these insects often cluster together, thread out, etc.
These are resistant to several antibiotics (pen, strep, Cipro, piperacillin, etc.), move extremely slowly, do not alter the color of the medium or cause it to become turbid, develop slowly under aerobic conditions, and may be found intracellularly as well as extracellularly. In addition, it has the ability to ruin any experiment. According to the findings of one study, this organism is not Achromobacter.
Also Trying an antifungal combination with kanamycin sometimes works, but it doesn't eradicate the problem.
I would ask that you please share your experiences with me so that something may be done.
Mohammad Mahdi Heidari Do these bugs you're talking about fall under the umbrella of "Mycoplasma" or are they something else unrelated?
Justin Clark
There is always a chance, but the most important fact to remember about mycoplasma is that its morphology is so small that it cannot be detected with a light microscope.
Even if we disregard this particular consideration, there is no way that they will be able to be painted using the DAPI technique.
We have this problem too, have you managed to get rid of the infection Khrisna Putera Buddhiman ?
Hello all, thanks for all suggestions and discussion Mohammad Mahdi Heidari Justin Clark . Rashmi Trivedi we didn't use a cell scraper. Rebecca Miller in the end, we weren't able to pinpoint the root cause (have tried changing the media as well), so we use the cell from another batch of source. Since the cell growth was slow, we were also thinking if this might come from a "cluster" of dead cell organelles.
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