SOP for immune monitoring of patient samples

immune monitoring of peripheral blood and bone marrow samples taken from the patients enrolled in the clinical trial described in this proposal are of utmost importance. Blood samples and if possible bone marrow samples will be drawn from the patients before the infusion of the multivirus- specific T-cell product and at different time-points afterwards. The presence of T cells directed against the targeted viruses (e.g. CMV, EBV and AdV) will be analysed by multi-colour flow cytometry using HLA/peptide multimers for known immunodominant viral epitopes combined with antibodies for different T-cell phenotypes (central memory, effector memory, memory stem T cells or effector T cells). Also restimulation experiments with viral peptide pools, followed by counterstaining with CD4/8 antibodies and staining for specific activation markers (CD137, CD154) and/or relevant cytokines (IFNγ, IL-4, GM-CSF) will be used.