What are the reasons for it and what type of precautions should be taken to prevent this type of morphology, the white granule appearance?
i have isolated human umbilical cord blood stem cell in P0 stage attaining confluency.....
i store the cells in CO2 incubator with 37degrees and 5% Co2
but the problem is initially they looked good but later on with increasing passage no they appeared shrinked.....can it because of the antibiotics or media or growth factors or is there any other reason ?
Yuetao Zhou ...thanks for your suggestion
but do you wanna say that they are seeded in wrong density ?
Its difficult to see from that picture but are the white granules in or on the cells? In primary cultures there will be some cells that initially attach, but then senesce and die producing cell debris.
To
Martin Stoddart, thanks a lot for your suggestion and taking out time to look in to the matter, but in the picture the white granules are present on the cells at a Passage number 3 and then they resided on P4 also and then the normal cells didnt grow ahead......
To, Purva Sanganeria.
According to me the reason could be:
1. Over-trypsinization: To prevent this reduce the concentration of trpsin/EDTA to its minimum prescribed concentration as well as duration of trypsinization. Secondly, wash your cells with either phosphate-buffered salin (PBS) or Hank' Balanced Salt Solution (HBSS) twice after trypsinization.
2. If you are using a low glucose medium (e.g., DMEM-LG), shift to high glucose medium (e.g., DMEM-HG).
[Reference: Deschepper M, Oudina K, David B, Myrtil V, Collet C, Bensidhoum M, Logeart-Avramoglou D, Petite H. (2011). Survival and function of mesenchymal stem cells (MSCs) depend on glucose to overcome exposure to long-term, severe and continuous hypoxia. J Cell Mol Med. 15(7):1505-14. doi: 10.1111/j.1582-4934.2010.01138.x.]
3. Increase percentage of FBS in the culture medium [I found 20% FBS works best for rat BMSCs]
I hope it helps.
Regards,
Fazal-ur-Rehman Bhatti.
Fazal-ur- Rehman , thanks a lot for your quality inputs
i will try them and will come back to you......
thanks a lot
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