Endothelial cell staining of Mouse Brain

[RYamada]

Hi all.
I'm currently trying to stain brains of murine xenograft models of glioma and having some difficulty getting a positive staining of the vasculature.

I've tried both frozen and paraffin sections with various antibodies including anti-CD31 (Invitrogen - 370700), anti-vWF (abcam - ab9378), and anti-Endoglin (Santa Cruz - sc-18838).

The protocol I use is pretty standard I think; with antigen retrieval for paraffin sections, using citrate buffer for 30 minutes etc.

I'm interested particularly in studying neovasculature.

Anyone with some experience in staining for mouse brain vasculature, I would really appreciate your advice on IHC protocol in order to get a positive staining.

Thank you.

R

[CanuckPhD]

I have used the following antibodies in acetone fixed frozen sections of rat spinal cord and brain. They worked quite well. However, it is known that CD31 is decreased in vessels that are inflammed during EAE (which was my study). Therefore I used collagen IV to quantify vessels. According to the datasheets the antibodies also work on mice (m in table below).

Good luck,

Collagen IV Rabbit polyclonal   r,m   Ab19808 2 μg/ml Abcam
CD31         Rabbit polyclonal   h,m   Ab78364 2 μg/ml Abcam

[MT Scientist]

We have used anti-eNOS (BD, monoclonal) and anti-vWF (Sigma, polyclonal) to stain ECs in sheep and rabbits with success.  I prefered the eNOS as the vWF always gave background.

Haven't have much luck with CD31 or CD34.

[JDenbeig]

Hi,

Can I ask if you ended up having success with your mouse brain staining, and if so, what concentrations for Endoglin you used?

Thanks
Janet