Hi everybody,
I am wondering if the unoccupied Ca2⁺ binding sites in soft alginate gel (gelation with low Ca2+ concentration) sequestrate the free Ca2+ of my cell culture medium. Which would also mean that the physical properties of the gel would not be stable over time, given that the culture medium is renewed every few days.
It seems that this issue is not discussed in the respective literature so I am grateful for your theoretical explanations as well as shared practical experiences in this matter.
Thank you, best regards
Jörg
Dear Panzer
The physical property of alginate hydrogel could be controlled and relatively stable by preparation conditions. Of course the free Ca2+ would be coordinate with COO- on alginate, meanwhile the coordinate COO-Ca2+ would release free Ca2+ becaues of ionization equilibrium. As a result, the physical property of alginate-Ca hydrogel would be relatively stable if the concentrations of free Ca2+,COO-Ca2+ and COO- kept relatively constant in several days.
Thank you Ton Zongrui!
So you mean when I change the medium after a few days, lets say 4, and with that increase free Ca2+ concentration, the gel parameters would not change because of the stable equilibrium during the first days?
Yes, I have found some papers which studied mechanical properties differences of alginate-Ca hydrogel incubated in PBS solution without Ca2+ ( doi.org/10.1080/09205063.2018.1433420) and CaCO3 solution (concentration 5.0 mM, doi.org/10.1002/jbm.a.31375 ) for different times.
In PBS, the modulus of alginate-Ca hydrogel would decrease gradually in 30 days. Meanwhile, the modulus of alginate-Ca hydrogel would increase by about 30% (in 5 mM Ca2+ solution) and then keep stable in 7 days. Consequently, the mechanical properties would be kept stable as long as you keep Ca2+ concentration stable even though you change the medium many times. (You could found in the second paper doi.org/10.1002/jbm.a.31375, they test properties of hydrogels with the medium changing every day)
Specifically, the Ca2+ concentration is 1.8 mM in a DMEM culture medium which could be found swelling ration changed barely in second reference (doi.org/10.1002/jbm.a.31375). It could be thought the mechanical of hydrogels would change barely because of the similar crosslink degree. Usually, the changing trends of swelling behavior and mechanical property were synchronously and could reflect the crosslinking degree.
Otherwise, if you worry about the varied mechanical properties of hydrogels in different mediums, you could keep the hydrogel in the cell culture medium you would use for one day to reach ionic equilibria.
Furthermore, how the mechanical properties of alginate hydrogels impact cell behavior or cell fate have been studied deeply by Dr. Chaudhuri (homepage: http://chaudhurilab.stanford.edu/index.html).
Some papers might be helpful to you:
Nam S, Stowers R, Lou J, Xia Y, and Chaudhuri O, Varying PEG density to control stress relaxation in alginate-PEG hydrogels for 3D cell culture studies, Biomaterials 200: 15 - 24 (2019). DOI: 10.1016/j.biomaterials.2019.02.004
Chaudhuri O*, Guo L*, Klumpers DD, Darnell M, Bencherif S, Weaver J, Huebsch N, Mooney DJ, Hydrogels with tunable stress relaxation regulate stem cell fate and activity, Nature Materials (2015). DOI: 10.1038/nmat4489
Branco da Cunha C, Klumpers DD, Li WA, Koshy ST, Weaver JC, Chaudhuri O, Granja PL, Mooney DJ. Influence of the stiffness of three-dimensional alginate/collagen-I interpenetrating networks on fibroblast biology, Biomaterials. 35(32):8927-36 (2014) DOI: 10.1016/j.biomaterials.2014.06.047
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