Hi
I need a good panel for my FACS experiment which I 'll differentiate (mature DCs- which pulsed with peptide) and immature DCs ?
Thanks
We routinely use CD11c, CD80, CD86 and MHC II for bone marrow-derived dendritic cells (BMDC). You just gate on CD11c positive cells and do mean fluorescent intensity (MFI) for all other markers. Immature DC express low levels of CD80, CD86 and MHC II where as mature DC express high levels of the same. Good luck
Usually a good BMDC prep has 90% of cells positive for CD11c after 6 days of colture. To check if cells are activated you can do an easy assay with LPS. You have untreated cells and treated for 24h with LPS and then you check the activation marker status (cd86, mhcII for example). They should be way higher in DCs treated with LPS than untreated. If results are similar, means that your DCs were previously activated. To prevent unspecific activation I suggest to be as gentle as possible when manipulating the cells.
Also you can see if cells are activated under the microscope...Activated cells stick to the plate and are bigger (usually) while resting DCs have a more circular shape and some of them will be barely attached to the plate.
Thank you So much Ramachandramouli Budida and Francesco Mainini helpful answers and tips:)
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