Could you separate the channels as follows:

actin/Hoechst

tubulin/Hoechst

Thanks!

Hi Nicholas,

We observed fair staining of nuclei and F-actin in the center of the spheroid whereas tubulin staining is mainly observed at the periphery. I don’t think the anti-tubulin antibody penetration is the problem as we performed ohter stainings on larger spheroid structure without any Ab penetration issue.

I would be interested in any paper/work dealing with tubulin expression in 3D spheroid.

Hi Adrien,

Would you be able to share your immunostaining protocol? I have some spheroids for a cancer cell line that i need to stain for various markers. 

Hi Adrien,

Could you please share with us your immunostaining protocol ?

We observed such features for actin in certain pancreas cancer cell lines as illustrated in the attached article.

Best regards

Robert