I am a beginner in cell culture and I would like to understand why aren't these MSC in collagen type I gel migrating? I kept the gels in DMEM+FBS 10% at 37°C/5% CO2. These captures were taken 1 month apart. Cells were stained with DiI.
What do you mean with the cells are migrating? Are they migrating during the culture period or after you stain with Dil? How many concentration of collagen type I that you use for the hydrogel fabrication? Also, did you add 0.5-1% antibiotics or not in the growth media? Please also explain in detail about the way you stain your samples for Dil staining. I wanna know first all the steps that you have been done before. Thank you.
Thank you!! I meant they are NOT showing any kind of displacement or movement, as I expected, but i don't know if maybe it is normal that there is no cell migration. I will explain the process:
I dissociate adherent cells from the flasks with trypsine, then I stained with DiI (5uL/10ml), I do this to stain them before seeding them in the hydrogel so I can "track them" when they are seeded. I let the DiI with the cells for 20 min in the incubator, then wash them with HBSS. I proceed to centrifugate and re-suspend in 1300 ul of collagen rat tail collagen type I EMD Millipore 08-115. Each scaffold or hydrogel was made with 300 uL cell suspension. Then I achieved collagen gelation using ammonia vapor chamber for 2-4 minutes. I only used 1% antibiotic once in the growth media because it was showing contamination but then stabilized. The hydrogels were kept in DME+10%FBS
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