Hi there,
I am new to the world of immuno-modulation and am looking for an in-depth protocol for the isolation of human peripheral blood mononuclear cells (PBMC)?

Does anyone have a protocol or know where to obtain one?

Thanks,
RockyDoc

Protocols will be there in most papers published in JImmunol or JLeukBiol that deal with human PBMCs - especially ones published ~8-10 yrs ago.
Basic protocols are also present in the "gold book" - Current Protocols in Immunology.
In brief, you use whole blood collected in Li-Hep or Hep-Cit "vacutainer" style tubes, subject it to Ficoll density gradient centrifugation (1.077) in order to obtain a "buffy coat". This can be further purified/enriched for any specific cell type that you intend to study - e.g. CD4+ T cells.

Here is an example of a typical density medium separation protocol:

http://www.cambrex.com/Content/Documents/Bioscience/Lymphocyte%20Separtion%20Medium.pdf

Also there is this paper:

http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=1749920&dopt=Abstract

This is also has some useful information:

http://www.beckman.com/literature/Bioresearch/A-1930Afinal.pdf

Thanks for the help!

We use Vacutainer CPT tubes. They are provided as citrate of heparine tubes. You should decide for what experiments you are going to use the cells.
After spinning you simply pipet of the PBMC fraction which is very clean. You do not need a laborious and cumbersome Ficoll method anymore. Our rsesults are very satisfying.
More information: www.bd.com

Hi there
we are trying to isolate PBMC from nude mice and using the BD vacutainer CPT. For quality control we tried staining with turk but we could get any positive staining. We are able to stain with tryphan blue and this worked but we still were questionable about what we are actually counting red or white blood cells? How did you make sure you had PBMC isolated? Did you use blood from Mice? Do you have a better method for isolation of PBMC from nude mice?