Comparison of Whole Blood Cryopreservation Methods for Extensive Flow Cytometry Immunophenotyping

Fresh blood immunophenotyping by flow cytometry, based on the reliable simultaneous detection of several markers in a cell, is the method of choice to study the circulating human immune system. Especially in large and multicenter studies, high sample quality is difficult to achieve, and adequate collection and storage of samples with fine-tuned whole blood cryopreservation is man-datory. Here, we compared the quality of immunophenotypic data obtained from fresh blood with those obtained after five cryopreservation methods by quantifying the levels of 41 immune cell pop-ulations. They comprised B and T lymphocyte subsets and their maturation stages, as well as mon-ocytes and granulocytes. Three methods used fixative solutions and two other methods used dime-thyl sulfoxide solutions to preserve cell viability. The fixative methods prevented detection of markers critical for identification of B and T cell subsets, including CD27, CXCR3, and CCR6. The other two methods permitted reliable discrimination of most immune-cell populations in thawed sam-ples, though some cell frequencies varied compared to the corresponding fresh sample. Of those two methods, the one preserving blood in media containing dimethyl sulfoxide produced results that were most similar to those with fresh samples.