3 replies to this topic

[netron]

Hi all,

Could someone share a protocol for preparing cells from fresch xenograft tumors, staining for a cell surface maker and then running FACS?

Thank you in advance.

N

[bcv85]

Hi, I tried a methods that worked for me with B16F10 melanoma.

I retrieved the tumor from subcutaneous and dissociated mechanically against a 70um cell strainer. From this cell suspension, counted the cells and made the dilutions (1x10⁶ cells/tube) for antibody staining.

In my case, I wanted to see leucocytes, then I stained cells with anti-CD45 and from CD45⁺ cells gate I analysed my populations.

I hope that was useful.

Bruno

[netron]

Thank you for the helpful information. Do you use any kind of enzyme to dissociate the cells besdies mechanic method? Can you store the tissue or cells for late analysis on FACS machine?

[pcrman]

I think the critical part is dissociating the tumor into single cells, the rest will be the same as cell lines. here is a very nice protocol for tumor processing and marker staining

http://sciencepark.m...essing4web.html