time required for cell recovery after thawing - (Jan/11/2008 )

Hi,

how long do you think cells (PBMCs, EBV transformed cells lines) should be incubated after thawing before proceeding with cell surface staining or functional assays?

Rana

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at least 48h but i would better go for 72h to efficiently let them recover functions and divide well.
The procedure says you have to plate them, split them once before use.

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I would give them atleast 2 days before trying any real experiment on them.

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Thanks!

For thawed primary PBMC you also allow 2 days recovery before non-functional assays such as surface staining? Do you know of a publication testing recovery of PBMCs? I would like to use the cells with minmal in vitro manipulation such as addition of cytokines therefore prefer using them as soon as may be appropriate.

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well this is not publication related but personnal and shared experience with colleagues and boss who deals with cells for a long time

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I am using human WBCs that have been frozen for ChIP. I compared using the cells straight after thawing, or after 24 hours in culture (seeded at 2 mill/ml). The ChIPs worked for both but I got much more DNA out of the cells that were cultured for 24 hour (just in RPMI, FCS, Glut). Hope this helps

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Thanks everyone.

@Clare: ChIP - that is DNA microarray?

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ChIP = chromatin immunoprecipitation And I use the DNA after ChIP to hybridise to an array (ChIP-on-chip).

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fast (<24h) dividing cell lines, after splitting, they start after 1 day with dividing which we take as the moment of good recovery and may begin with transfection...

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