is a matrigel layer a 3d cell culture? - (Feb/28/2008 )

thats my question:

i know that 3d cell culture is achieved with sponges made out of alginate or similar marerials.
i never worked with matrigel but the way i understood it you pipet the matrigel suspension in your well and the proteins in it will self-assemble into a sort of framework for your cells to grow on. is that correct?
so my question is: is this framework similar to an alginate sponge for 3d cell culture?

there was a thread some weeks ago about how to make the matrigel layer as thin as possible. so on the contrary: how high can you stack matrigel up?

thanks in advance

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No, its not similar to 3D cell culture. Matrigel has substrates which provide good cell adhesion and growth. But I believe it could be used in a 3D cell culture system.

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Hi
have one of you already done histochemical staining of in matrigel grown cells? I ask because I want to show definitely a postive localization of my protein in the cell nucleus. A confocal laser scan through cells which were grown on coverslips are not satisfying. Any idea?
Tiffy

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yeh, we regularly do immunostaining on cells grown on matrigel coated coverslips.

If the staining is not working check antibody. Also you could grow them without matrigel and look at them

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scolix specultes right, matrigel can be used for 3D culture but it will be expensive; think of collagen instead

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The antibody works very well. It would be very kind if you describe how you stain your cells which were grown in matrigel. I have no experience. How do you fix your cells? PFA? Which detergent do you use to permeablize your cells. And how do you embed or mount your cells. Where are the pitfalls of this staining.
Thanks a lot
Tiffy

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We wash cells with PBS, fix with 4% PFA(10min), wash PBS, permeabilize with triton-0.2% (5min),wash PBS, blocking, primary antibody, 3x wash-PBS, secondary antibody, 3x wash-PBS, DIP in water-30seconds, mount it (vectashield).

Good Luck !!!

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