Monocyte isolation from mice - (Sep/26/2008 )

I am new to monocyte work and I would like to get a protocol which describes the best way to isolate enough mouse monocytes to be able to make automated planar patchclamp recordings from on the day of isolation and even maybe to try and culture them for a week or more. Isolation from whole blood will not give me enough cells (1 million/ml required) for this and someone suggested peritoneal lavage or extraction from the spleen. I would also like to know if anybody does this kind of preparation in the UK where I might come and learn first hand. CJM

hi,
collect the spleen from mice, leave it in 5-10ml of DMEM alone depending on the dish u use under sterile conditions. make a little cut at one end of the spleen using a sterile foreceps,
take DMEM into the 5ml syringe, plunge the medium through the spleen.
you will observe dark red in ur DMEM, those are splenocytes.
repeat this step 2-3 times.

spin down cells, if you want kill RBC or proceed with other steps.

gud luk