Core B: Cell Banking and Immune Assessment Core

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Project Summary/Abstract Core B The purpose of this core is to provide a centralized laboratory resource for obtaining, cryopreserving and distributing patient samples and to monitor the kinetics of immune reconstitution following allogeneic hematopoietic stem cell transplantation (HCT). Prior to transplantation, peripheral blood mononuclear cells (PBMC), bone marrow and serum or plasma are obtained from all patients enrolled on clinical trials supported by this Program Project. PBMC, plasma and serum are also obtained from normal stem cell donors for these individuals. Following HCT, peripheral blood, plasma and DNA samples are obtained at regular intervals. All of these samples are processed, cryopreserved, entered into a computerized inventory and made available to all of the investigators in this program. To support clinical trials in Project 1, the effects of immunologic interventions are evaluated through serial assessment of specific lymphocyte populations that define changes in number as well as their level of activation, maturation and other functional characteristics. Phenotypic analysis of circulating lymphocytes primarily utilizes multi-parameter flow cytometry with a panel of fluorochrome-conjugated monoclonal antibodies. Plasma concentrations of cytokines known to play important roles in immune reconstitution (BAFF, IL-2) are measured by ELISA. Reconstitution of B and T cell receptor repertoire is examined by in depth sequencing of immunoglobulin and TCRV genes. Thymic function is evaluated by quantitative PCR measurement of T-cell receptor excision circles (TREC). Reconstitution of T cell immunity to specific target antigens such as CMV and EBV is determined by ELISPOT and HLA-tetramers. In Project 3, patient plasma is used to identify antibodies specific for HY antigens or other allo or auto antigens. Results of each of these assays are correlated with other parameters of immune function as well as with clinical outcomes.