Hi all,
I am having trouble with collagen type I as a scaffold for cells.
I have a 3mg/mL collagen I solution with pH = 2. I am neutralizing with 1 M NaOH to pH ~7, then mixing with ice cold cell suspension (with DMEM/F12 and supplements) to 1,5 or 2,5 mg/mL collagen. I seed the collagen/cell suspension into chamber slides (cell culture treated polymer bottom) and after 1 or 2 h at 37°C I cover it with medium. I observe that the hydrogel is not stable as it degrades on the macroscopic level. Parts of can be seen floating around, rest seems to dissolve within days. When I wash for doing immunostaining, hardly anything of the gel remains attached.
I feel that when working with high(er) collagen concentrations, the gel is more stable. But for the neurons the stiffness must be low for them to grow. And I know that other groups worked with even lower collagen concentrations.
Does anybody has experienced similar problems or knows how to solve the problem?
Thank you,
Jörg
I guess the stability of the gels is being disrupted by the high salt concentration of cell culture media. I have had similar problems with other gels. Try to cross-link the gel for increasing their stability. You might have to try a few methods for stiffness optimization.
Thank you for your comment Nabodita Sinha.
In the initial gel, I have only 1/6 of the volume as culture medium. I reckon the salt concentration is not that high then. Is there a method for cross linking that you can recommend?
Cheers
Cross-linked Collagen Hydrogel Matrix Resisting Contraction To Facilitate Full-Thickness Skin Equivalents
This paper might serve as an example
- Badges
- Science topic
- Similar topics
- Gels