>In my intracellular staining protocol, PMA/ionomycin stimulation >is usually done the day after blood collection and PBMC ficoll >separation. Does anyone know what is the best overnight storage >temperature for PBMC? I did 37 °C, 5 % CO2 and also closed tubes at >4°C, both in RPMI supplemented with BCS. For certain cytokines these >2 conditions really give different results! We store PBMC overnight, but usually are working with mobilized cytapheresis products, and then performing either CD34 selection, or initiating cell expansion/transduction protocols. I can't, therefore, say whether our storage methods will be suitable pre-PMA/ionomycin stimulation, but perhaps the information will be of some help. In our experiments with pre-processing cell storage, we obtained slightly better viability and viable mononuclear cell recovery when cells were stored at 4oC in gas-permeable bags. Ambient temperature was almost as good, however. We did not test 37oC. We obtain very good results storing cells in a solution called STM-Sav. This may not be commercially available where you are located, but you can make a good facsimile from lactated Ringer's, your buffer of choice, and 1% albumin. You might take a look at this paper for further information: Burger SR, Hubel AH, McCullough J. 1999. Development of an infusible-grade solution for non-cryopreserved hematopoietic cell storage. Cytotherapy 1:123-133 Scott -------------------------------------------------- Scott R. Burger, M.D. Director, Cell Therapy Clinical Laboratory Assistant Medical Director, Blood Bank University of Minnesota, Fairview-University Medical Center 612-626-4919 612-625-2199 - Fax burge009@tc.umn.edu http://www.mbbnet.umn.edu/labs/ctcl.html http://www.borg.labmed.umn.edu/faculty/burger_s.html
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