Dear colleagues!
We are facing several problems while trying to establish primary rat endotheliocyte cultures.
First of all, cells loose viability after cryoconservation. The freezing medium used is 90% FBS supplemented with 10% DMSO. After thawing there are just no live cells in sight, only some debris floating around. What could be an issue? Do these cells require some special treatment?
Next we are failing to grow the cells on coverslips for subsequent ICC analysis. Tried coating glass with collagen, laminin or polyornithine. The cells don't adhere no matter what. Some reserchers mention fibronectin in their papers as a proper coating agent, but we currently don't have any in posession. Is there any chance fibronectin may help?
Any comments and recommendations will be much appreciated!
Yours,
George Leonov
Laboratory of Cell Biology
V.N. Orekhovich Institute of Biomedical Chemistry, Moscow, Russia
Hello Georg,
Please see the attached article for growing primary rat brain ECs.
Dear George,
For freezing, use CryoSFM from PromoCell. It really makes a difference, but do not keep cells in liquid N2 for more than 1 year.
Regarding coating, a combination of collagen type IV and fibronectin is the best.
Hope I could help.
Best regards,
Imola
- Badges
- Science topic
- Similar topics
- Engineering
- Materials Engineering
- Coating