Assuming the use of FCS+10% DMSO as cryoprotectant and 1ml of final volume.
I believe these two manuscripts might be of assistance
Disis ML, dela Rosa C, Goodell V, et al. Maximizing the retention of antigen specific lymphocyte function after cryopreservation. J Immunol Methods. 2006;308:13–18
Mallone R et al. Isolation and preservation of peripheral blood mononuclear cells for analysis of islet antigen-reactive T cell responses: position statement of the T-Cell Workshop Committee of the Immunology of Diabetes Society. Clin Exp Immunol 2011 Jan 163(1) 33-49
("no significant differences in viability were observed when PBMCs were frozen at concentrations of 1 × 107, 2 × 107 or 3 × 107 cells/ml per cryovial (1 ml per cryovial). Thus, it seems safe to recommend freezing PBMCs at concentrations ≤ 3 × 107 PBMCs/ml (level of evidence: E). Higher concentrations have not been tested systematically ")
Usually, 1 million / ml, in 10%DMSO, 40% FCS and 50% complete medium.
We routinely freeze 1x10^7 in 1ml of 90% FBS / 10% DMSO, and recover about 95% viable cells when thawing. You could try 5x10^7, but your recovery might suffer a bit.
I believe these two manuscripts might be of assistance
Disis ML, dela Rosa C, Goodell V, et al. Maximizing the retention of antigen specific lymphocyte function after cryopreservation. J Immunol Methods. 2006;308:13–18
Mallone R et al. Isolation and preservation of peripheral blood mononuclear cells for analysis of islet antigen-reactive T cell responses: position statement of the T-Cell Workshop Committee of the Immunology of Diabetes Society. Clin Exp Immunol 2011 Jan 163(1) 33-49
("no significant differences in viability were observed when PBMCs were frozen at concentrations of 1 × 107, 2 × 107 or 3 × 107 cells/ml per cryovial (1 ml per cryovial). Thus, it seems safe to recommend freezing PBMCs at concentrations ≤ 3 × 107 PBMCs/ml (level of evidence: E). Higher concentrations have not been tested systematically ")
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