Dendritic cell novice - (Nov/16/2005 )

OK, I am a dendritic cell novice, and actually an Immunology novice.

i want to know:

1-generating DC from monocytes or BM: Has anybody used Human serum in the culture (rather than FCS described in all) and whats the results? If you are looking at maturation and more specifically of the effect of an hormone on it, what time points should you be looking at after the "maturation agen" ( LPS or TNF) has been added.

2- If you wanna look at DCs migration or infiltration induced by cytokines: has anybody got any protocols in vitro or in vivo, and do you need to add an antigen somewhere?

Thanks
Bouka

forgot:
has anybody got any experience of labelling DCs with CSFE. if yes, how long do the cells stay fluorescent in vitro and in vivo ( ie when you should detect them).

I use FCS, and usually have good results.
for maturation, add LPS or TNF-a for 24-48 hours.
CFSE labelling works well with DCs, but CFSE bleaches a little. The main problem is that when you inject DCs in vivo, only a small fraction reaches the spleen or LNs. May be adding a maturation signal in vivo would help the migration.

thanks##


quote name='mg2g2h' date='Dec 6 2005, 09:17 AM' post='33637']
I use FCS, and usually have good results.
for maturation, add LPS or TNF-a for 24-48 hours.
CFSE labelling works well with DCs, but CFSE bleaches a little. The main problem is that when you inject DCs in vivo, only a small fraction reaches the spleen or LNs. May be adding a maturation signal in vivo would help the migration.
[/quote]