dendritic cells culture - (Nov/16/2007 )
How to culture dendritic cell from monocytes CD 14 which is isolated from PBMCs by MACS? thanks
I am still confuse, how much GM-CSF and IL-4 concentration? I followed procedure from journal (used GM-CSF 500 IU/ml and IL-4 250 IU/ml), but it doesn't work. I thought the monocytes changed to macrophage not immature dendritic cells.
-m.anggelia-
QUOTE (m.anggelia @ Nov 16 2007, 01:35 AM)
How to culture dendritic cell from monocytes CD 14 which is isolated from PBMCs by MACS? thanks
I am still confuse, how much GM-CSF and IL-4 concentration? I followed procedure from journal (used GM-CSF 500 IU/ml and IL-4 250 IU/ml), but it doesn't work. I thought the monocytes changed to macrophage not immature dendritic cells.
I am still confuse, how much GM-CSF and IL-4 concentration? I followed procedure from journal (used GM-CSF 500 IU/ml and IL-4 250 IU/ml), but it doesn't work. I thought the monocytes changed to macrophage not immature dendritic cells.
Why would a scientist put correct information, that they got from hard work, in a journal so everyone could get it for free???
M-CSF will push monocytes into macrophages and not DCs. Give me the rest of your protocol.
-dave2-
This is the protocol I used on PhD. Can seem to find the cell concentration though 
PBMC were purified by CD14 positive selection of Buffy coats. Cells were incubated with 80 ng/ml huGM-CSF and 500 U/ml huIL-4 (Biosource) in 75 cm2 flasks for 6 days. Media wash replaced every 3 days, and on day 6 cells were replated at the appropriate concentration for treatment.
hope it helps some.
-almost a doctor-
QUOTE (almost a doctor @ Nov 20 2007, 03:38 AM)
This is the protocol I used on PhD. Can seem to find the cell concentration though
PBMC were purified by CD14 positive selection of Buffy coats. Cells were incubated with 80 ng/ml huGM-CSF and 500 U/ml huIL-4 (Biosource) in 75 cm2 flasks for 6 days. Media wash replaced every 3 days, and on day 6 cells were replated at the appropriate concentration for treatment.
hope it helps some.
PBMC were purified by CD14 positive selection of Buffy coats. Cells were incubated with 80 ng/ml huGM-CSF and 500 U/ml huIL-4 (Biosource) in 75 cm2 flasks for 6 days. Media wash replaced every 3 days, and on day 6 cells were replated at the appropriate concentration for treatment.
hope it helps some.
This means you used .08mcg/ml of GM-CSF which is a little less than 500IU/ml. Is this correct?
-dave2-