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DC harvesting for FACS - dendritic cells (Jul/24/2005 )

I was just wondering how everyone harvests their DCs after stimulation for FACS. We have done it with either scrapping where we kill alot of cells or with trypsin but of course we either lose cells due death by scrapping or lose magnitude og signal due to the trypsin. Does anyone have a different method of harvesting that doesn't have these draw backs?


aspire the culture supernatant
add few mL of PBS/EDTA 2mM for rinsing and recover the PBS/EDTA
add few mL of PBS/EDTA and let "unhook" at RT for 5-10 min
recover this suspension, and so beat (slap or bang in English?) the dry plate for harvesting the maximum of cells you can resuspend in the later supernatant before a centrifugation


we "scratch", quite effectively