Protocol for mouse splenocytes - storage, media etc (Apr/13/2005 )

Hi

I am new in cellular immunology and therefore I really need help.

Do anyone have a working protocol for the isolation of leucocytes from mouse spleen.

I would like to get details of the media composition, hints and other things "that you just need to know".

Can the splenocytes be stored for 2 days before use, and if so, do they need som special media or do you just change the medium daily.

I expect to use the cells for ELISPOT.

Thank you very much in advance

The novice

we use RPMI 1640 supplemented with 5% FCS and 10mM HEPES for our mouse splenocytes, medium is changed every day, but don't expect them to be very longlived... 1 Week is possible, though....

splenocytes are isolated by grinding the spleen on a cell strainer (100µm) and lysing the erys... if you want lymphos only, you'll have to do depletion of the other cell types or positive selection of lymphos by magentic beads, for example...

mike

Thank you very much for the fast reply.

Cheers

J