I wish to use frozen leucocytes (PBMCs purified on ficoll column) -how long wake-up time do they need ? (incubation time prior to stimulation in the ELISpot plate)

Hello,



	After thawing frozen PBMC we usually rest them for 1h in the incubator before proceeding with our ELISpot/FluoroSpot assay. During the 1h incubation many apoptotic cells will tangle up and die. A white cell "gew" is formed in the 15ml tube. As a result, after the 1h incubation I always transfer the living cells to a new 15ml tube and leave the cell-clumps and gew in the previous tube. 



	I then setup my ELISpot.

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Frozen, cryo-preserved cells