What passage are the cells? What media are you using. In the first two photos, lots of small "debris" may be indication of contamination.
Hello as others before me have stated, it is kind of hard to help without knowing specific culture conditions (passage, medium, plastic type or coating etc..)
Hello
I agree with scientists . Cells are not too bad , also you have contamination clear during culture . pleas , make sure of the validity of the reagents and materials used during culture " If approached on expiration date would prefer not to use them " . also , told us about the protocol that you used so that we can help you .
Best wishes
Third image shows some nice cells that look like stem cells but its hard to tell why culture ends up looking like other images. Feeding medium; contamination; other cells in culture? Its hard to know without more information.
Thank you all for your feedbacks
I have been using StemGro hMSC Growth Media from life tech.
The passage numbers are either 4 or 5.
I was expecting more elongated fibroblast-like shape but somehow, I am getting this
Hi
I would suggest taking a look at this paper: (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2707099/pdf/joa0214-0759.pdf),
In Fig.1 they show MSC with different characteristics, and to me your cells look more like the ones labeled with 'SS', which are slowly dividing cells, and more tend to differentiate into cartilage yet are still MSC. Depending on the donor (assuming your cells are human MSC) they show different characteristics, one sample might rapidly differentiate into adipocytes, another one might be proliferating faster, etc. I would suggest keeping the ones you already have in culture, and also start a new culture from a different donor if you can to see if it is cell dependent or you need to change culture conditions.
You have a mixture of cells present in your cultures, not just mesenchymal stem cells (Thy-1, CD90, CD13+). You also have "contaminating" mesenchymal progenitor cells (CD105, CD123, CD166+), pluripotent stem cells (SSEA+), totipotent stem cells (CEA+), and fibroblasts. My suspicion is that your are using a serum that has not been pre-tested for inductive agents. Suggest using a heat-inactivated serum for better results. You also appear to be growing the cells on a substrate other than type-I collagen, a pre-requisite for mesenchymal stem cells. Suggest coating your dishes with a solution or 1% w/v type-I collagen in DPBS (easiest to get into solution by autoclaving on wet cycle, also sterilizes at same time. Coat surface, allow to air dry, lasts about 1 year stored at room temperature) or purchasing Falcon plasticware that has been pre-coated with porcine type-I collagen.
Hi Chi Woon Yoon. Just my two pence. I agree with others that your cells look fine and especially as Dr. Young has kindly pointed out, there suspiciously looks like a mixture of cells. Along with all the suggestions, if time permits you, you may try using different passaging solutions (when next culturing earlier passages). As MSCs are highly adherent cell types, a mild passaging solution like accutase might help you to remove other cells quicker. So lesser time with the passaging solution and mostly the MSCs get left behind. But always cell sorting (if you have the option) is a better way to go. Have a good day.
I would say that either these cultures are contaminated with a lot of non-MSCs (ie, fibroblasts, endothelial cells, etc) or that they have become senescent. You could run them through a flow cytometer to check expression of non-MSC markers, such as the ones Dr. Young suggested.
Greetings Chi Woo,
There seems to be a 'contamination' on the images you've uploaded, especially those debris. Also encountered these on few of our primary lines. However, I would shoot one question that might be far from the topic, but: What would you pick between primary mesenchymal stem cells and commercial cell lines and why? As a general fact.
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