Umbilical cord blood

[Vos nom et prénom]

I give you my methodology of separation of cord blood cells :

- Dilute anticoagulated cord blood 1:4 with PBS-5mMEDTA and overlay 35 ml
on 15 ml of Ficoll.
- Spin for 35' at 400 x g.
- Collect interfaces and wash two times in PBS/EDTA. Centrifuge at 200 x g
for 10'.

The dilution is very important in this protocole.

Good luck. 

Florence

Doris Anne Morgan <dormor at bellatlantic.net> a écrit dans l'article
<353806C4.F2F3F998 at bellatlantic.net>...
> As anyone recovering the CD34+ cells from cord blood knows from
> experience, RBC contamination is very frustrating and overwhelming.
> Does anyone have an idea as to why the red cells of the neonate do not
> pellet in Ficoll? Is it a density difference of the newly formed red
> cells? Also I have found them to be quite resistant to hemolysis.
> Membrane difference? If anyone is routinely working with cord bloods,
> would really enjoy sharing some of these technical problems.
> 
>