I am trying to maintain the cell confluency around 90-100% in the glass culture plate. but cells are changing their morphology into spheroid-like structures by shrinking cells and are likely to be differentiated into other cells type (not sure).
and if I seed the cell at 20000/cm2, on the following day cells begin to devolve this morphology, whenever the cell confluency is less than 50%.
I want to maintain a monolayer of cells at high confluency on a glass culture dish.
How to solve this problem??
Please look at the attachment and give suggestion
Surely, because of high proliferation and lack of space, cells are peeling off. Well, could you please share the media you are using, the percentage of FBS, and the passage numbers?
I am using a Mesenchymal Stem Cell Basal Medium (ATCC- PCS-500-030), and Mesenchymal Stem Cell Growth Kit for Adipose and Umbilical-derived MSCs - Low Serum (PCS-500-040). and passage no is 5-6. I think cells are not peeling up, they are aggregated and develop into a spheroid-like structure.
I suggest using coating substrate to enhance the attachment of UC-MSCs on the glass culture plate. Vitronectin or CellStart might be the one.
I observed sometimes when cells get over-confluent then they peel off like this and the rest attached cells form a cluster. Also, during isolation, after WJ digestion when I plate the digested pellet, the next day, I see a round cluster attached at different locations of the plate and with each passing day, cells come out from those clusters and spread. After 10-12 days, clusters disappear and I get a homogenous monolayer of WJMSCs. Well, you mentioned a glass culture plate, but I never used it. Does, the plates coated or not? if not you may need a coating or coated plate to overcome the situation.
Duc M. Hoang and Saurabh Mandal thank you so much for the response.
I am using 0.1% Gelatin Solution for coating. Duc M. Hoang, Does Vitronectin or CellStart will work better ? I am trying to differentiate UCMSC, so Is it okay to use Vitronectin or CellStart?
Mahesh Prakash Bhatta I suggest you use cell culture treated plate (Nuclon Delta surface) for MSC differentiation. If you use culture treated plate, you dont have to coat as MSCs will attach firmly to the plate. In case you wanna use glass surface culture dish, I would recommend using Vitronectin or CellStart for differentiation.
- You could have some ideas from our paper: Standardized xeno- and serum-free culture platform enables large-scale expansion of high-quality mesenchymal stem/stromal cells from perinatal and adult tissue sources (10.1016/j.jcyt.2020.09.004)
- Badges
- Science topic
- Similar topics
- Biological Science
- Anatomy
- Tissue