A-098 Preliminary Tests of Cell Viability, Proteins and mRNA after Freezing PBMC for the Study of Chronic and Acute Lymphoproliferative Leukemias

Abstract The peripheral blood mononuclear cells (PBMC) cryopreserved are widely used in human immunology. However, PBMC, like other cell types, are sensitive to the freezing process. The impact of cryopreservation on T and B cell subsets of PBMC remains a challenge. For human PBMC, some reports have not observed significant changes between fresh and frozen cells in terms of proportions, phenotype and functionality, as well as antigen-specific responses or response to mitogens. Others, however, report that cryopreservation alters the proportions of human PBMC subsets in various immune responses. With the interest of studying chronic and acute lymphoproliferative leukemias (CLL and ALL), the authors’ objective was to test the viability of PBMC cells obtained from patients with CLL and ALL before and after freezing, following a simplified protocol. Methods On blood samples collected in EDTA, readings and cell counts were performed. Samples with a suggestive diagnosis of CLL and ALL were selected for the separation of PBMC whose analyzes were performed before and after freezing. The freezing of the PBMC was carried out in several stages and the whole process is carried out with care and in a gradual and simplified way of freezing until reaching the liquid nitrogen phase. Cells were kept frozen in freezing medium (RPMI plus 3% DMSO and 20% fetal bovine serum) for 1 week. Afterwards, staining with Trypan blue and counting in a Neubauer chamber were carried out. Student’s t test for independent variables and a significance level of 5% were considered. Results Table 1. Conclusion The description of this technique allows its use for analysis of gene expression profile, being able to contribute with the description of the molecular signature of leukemias.