This picture is taken after a one-week culture in droplet. Why do the chondrocytes become like this? I guess maybe it's due to a lack of nutrients or the change of osmotic pressure. What do you think? Looking forward to your suggestions and responses.
Thanks!
What your are trying to achieve. What do you mean by fibrosis in vitro?
I did work on nucleus pulpous progenitor cells, they made very beautiful spheres.
I used to filter cells through 70 strainer to make it single cell suspension and plate in a low-adhesion flasks (important) at a density of 10cells/ul media (100, 000 cells/10ml). After spheres become very big after many days, they tend to adhere to bottom. Before they adhere, to expand them, strain whole suspension of spheres thru 40um strainer, collect the spheres by reverse washing the filter and digest with enzyme and replate them for second round of culture.
Hope that will help
It is a great help. I wanna get a great amount of cell microspheres. I think this way can be a very good candidate, because my way, the hanging drop culture, is pretty expensive. What's the worst shortcoming is that you have to use many plates to gain enough microspheres, because in single drop there is only one or maybe two microspheres. Thank you for you suggestion, At the beginning of graduate life, it is a little difficult to do something, especially in the field that has nothing to do with your major as a undergraduate student.
This happens when the adhesion between the cells is greater than their adhesion to the surface they are grown on. You can induce it with low serum or growing on a low adherence surface or by generally making the cells unhappy. Chondrocytes seem to be very good at making clumps whereas other cell types will lift off plates as whole sheets. The only problem with tight balls of chondrocytes produced like this is that, depending on their size, they quickly become necrotic in the centre as the nutrients can't diffuse through the tight cell mass so we prefer to use them in a 3D culture system such as alginate or a scaffold.
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