Method for quantifying MSCs in a digested umbilical cord

[Maddy78]

Hi
I have a Beckman Coulter FC500 and am interested in determining the number of mesenchymal stem cells in a mixed population from a digested umbilical cord. Any ideas on setting up a protocol to do this and markers/ controls etc. Also what gating to use?
Thanks

[Bonnie]

In my opinion, the defining paper on MSC surface markers is:

Cytotherapy. 2006;8(4):315-7.
Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement.
Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D, Deans R, Keating A, Prockop Dj, Horwitz E.

That paper says that MSCs are defined as positive for CD105, CD73 and CD90, and lack expression of CD45, CD34, CD14 or CD11b, CD79alpha or CD19, and HLA-DR. If you set up a multicolor experiment to test all or most of those (you can probably get by with 105, 73, 90, 45, and 34), any reviewer will agree that you're correctly identifying MSCs.

As a control, you should get some MSCs that you know are good (ie. from Lonza or wherever) and verify that they show the correct phenotype in your experiment. You'll also want something positive and something negative for each marker in order to set your FACS gates; if you have lots of miscellaneous cell lines available in your lab, you can probably find ones that express each of the markers without buying any new cells.

[Maddy78]

Hi Bonnie
Thanks for the advice. Paper looks useful too.

[Maddy78]

Hi Bonnie
Any idea where I can get access to this article for free- I don't really want to subscribe for one paper.
Thanks