Journal: Cells

Article Title: Serial Measurements of Apoptotic Cell Numbers Provide Better Acceptance Criterion for PBMC Quality than a Single Measurement Prior to the T Cell Assay

doi: 10.3390/cells4010040

Figure Lengend Snippet: Percentage of live, dead, and apoptotic T cells, and T cell functionality after storing PBMC at 4 °C or 37 °C overnight. PBMC were stored at 4 °C or 37 °C overnight. ( A ) The numbers of live, dead, and apoptotic cells was determined after overnight storage. ( B ) PBMC stored overnight at 4 °C (solid bars) and 37 °C (open bars) were resuspended in fresh medium and tested in an IFN-γ ELISPOT assay with different antigens. CMV peptide pool, EBV peptide pool, and Flu peptide pool were used to activate CD8 cells and Mumps and Mosquito antigens to activate CD4 cells.

Article Snippet: The antigens CEF peptide pool (CTL-CEF-002), CMV pool (PA-CMV-001), Flu pool (PA-Flu-001), and EBV pool (PA-EBV-001) were obtained from Cellular Technology Ltd., OH USA.

Techniques: Enzyme-linked Immunospot

Journal: Clinical & translational oncology : official publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico

Article Title: Evaluation of safety and efficacy of p53MVA vaccine combined with pembrolizumab in patients with advanced solid cancers

doi: 10.1007/s12094-018-1932-2

Figure Lengend Snippet: p53MVA/pembrolizumab activate p53-specific T cell responses and associated immune function pathways. The response of CD8+ and CD4+ T cells from PBMCs after 24-h stimulation culture with p53MVA, MVA, p5396, and pp65138, as determined by flow cytometric analysis, is shown for five patients in columns a and b. The upregulation of CD137 expression on the surface of T cells in response to specific recall stimuli reflects increased frequencies of p53-reactive T cells in the circulation after vaccination. Culture conditions: NIL – medium alone; p53(96) – pool of peptides derived from wild type p53 sequence; pp65(138) – control peptides derived from pp65 CMV; MVA – wild type MVA vaccinia virus; p53MVA – recombinant MVA virus. Column c shows CD4+/CD8+ T cell ratio data for patients UPN003, UPN008, and UPN006 with declining ratio who benefited from the treatment and two patients UPN002 and UPN004 (and all other patients in Fig. 2) whose ratio remained stable and unaffected by the treatment. Column d summarizes data from multiplexed gene expression analysis of PBMC samples from indicated patients using nCounter PanCancer Immune Profiling Panel. The analysis of 730 immune profiling genes included selected genes that define immune function pathways. The pathway scores are plotted to show how they vary across time during treatment. The T cell functions and associated immune response categories remained at elevated levels for prolonged period of time in 2/3 patients responding to the treatment. Data presented for UPN003 in columns a, b, and d have been modified with additional time points that were not available at the time of their original publication in Yuan et al. [15].

Article Snippet: In the initial analysis, PBMC were thawed and plated at 2 x 10 5 cells/0.2 ml/well in media (RPMI, FBS 10%, glutamine 2 mM, sodium pyruvate 1 mM, non-essential amino acids) with one of the following stimuli: media alone (NIL), p53MVA, MVA, pool of 96 15-mer overlapping peptides spanning the entire length of p53 (p53 96 ; 5 μg/ml; synthesized in-house), and a control pool of 138 peptides derived from CMV pp65 protein epitopes (pp65 138 ; 2 μg/ml; BEI Resources, NIH, Bethesda).

Techniques: Expressing, Derivative Assay, Sequencing, Control, Virus, Recombinant, Gene Expression, Modification