positive cd3 cell selection kit ii  (STEMCELL Technologies Inc)

Journal: Science Advances

Article Title: Exposure of volunteers to microgravity by dry immersion bed over 21 days results in gene expression changes and adaptation of T cells

doi: 10.1126/sciadv.adg1610

Figure Lengend Snippet: ( A ) Schematic representation describes the experimental procedure. Blood was taken from eight healthy volunteers at five time points: 7 days before DI exposure (day 0, orange); 7 (blue), 14 (teal), and 21 (green) days during DI exposure; and 7 days after DI exposure (day 28, pink). CD3 + T cells were isolated from the blood, and bulk RNA-seq was performed. ( B ) Horizontal bars show the fraction of granulocytes, lymphocytes, and monocytes in blood isolates before CD3 + T cell isolation as determined by flow cytometry. Values are indicated as means + 1.96 × SEM. ( C ) Horizontal bars display the estimated fraction of T cell populations detectable in the bulk RNA-seq data as determined by deconvolution analysis with the quanTIseq algorithm. Values are indicated as means + 1.96 × SEM. ( D ) Factorial map of the PC analysis separates bulk RNA-seq gene expression data of each individual volunteers (small dots) and mean values across all volunteers (large dots). The proportion of variance explained by each PC is indicated in parentheses. Color-coding is according to the different time points. ( E ) The dot plot shows PC1 separation of bulk RNA-seq samples by time point. Individual values (small dots) and mean values (large dots) are shown per time point (color-coded). Error bars indicate means ± 1.96 × SEM. ( F ) Illustration displays differentially expressed genes (DEGs) identified by pairwise time point comparisons using DESeq2. The number of significantly (adjusted P < 0.01) up- [positive log 2 FC (fold change), red] and down-regulated (negative log 2 FC, blue) genes is indicated. ( G ) Line charts show the two k -means clusters and number of differentially expressed genes per cluster detected in at least one time point comparison. Standardized gene expression profiles for individual genes (gray lines) and the cluster center (connected dots) are shown.

Article Snippet: Human T cells were enriched by the positive CD3 cell selection Kit (EasySep Human CD3 Positive Selection Kit II, STEMCELL Technology) with a purity of 90 to 95%.

Techniques: Isolation, RNA Sequencing, Cell Isolation, Flow Cytometry, Gene Expression, Comparison

Journal: Science Advances

Article Title: Exposure of volunteers to microgravity by dry immersion bed over 21 days results in gene expression changes and adaptation of T cells

doi: 10.1126/sciadv.adg1610

Figure Lengend Snippet: ( A ) Heatmap demonstrates expression of overall down-regulated genes in CD3 + T cells identified upon DI (cluster 3 in fig. S9, n = 148) (left ) that overlap with genes in CD4 + (middle) and CD8 + (right) T cells obtained in the NASA twin study. Gene expression changes are shown as log 2 FC to day 0 for DI and log 2 FC for relevant comparisons in the NASA study (gray if missing). Relevant genes are highlighted. ( B ) Vertical bars display changes in expression for selected genes [ x axis annotated according to (A)]. Log 2 FC values show up- (red) and down-regulation (blue). ( C ) The dot plot denotes mean fluorescence intensity (MFI) ratio of cell surface marker CD25 compared to control as determined by flow cytometry. Values for individual volunteers (small dots) and mean values across all volunteers (large dots) are shown for each time point (color-coded). Error bars indicate means ± 1.96 × SEM.

Article Snippet: Human T cells were enriched by the positive CD3 cell selection Kit (EasySep Human CD3 Positive Selection Kit II, STEMCELL Technology) with a purity of 90 to 95%.

Techniques: Expressing, Gene Expression, Fluorescence, Marker, Control, Flow Cytometry

Journal: Cancer immunology, immunotherapy : CII

Article Title: Absence of host NF-κΒ p50 induces murine glioblastoma tumor regression, increases survival, and decreases T cell induction of tumor-associated macrophage M2 polarization

doi: 10.1007/s00262-018-2184-2

Figure Lengend Snippet: Gliomas in p50−/− hosts have increased total and activated CD4 T cells and, to a lesser extent, increased activated CD8 T cells. a CD4 or CD8 cells were quantified as total number per tumor and the relative number per tumor bioluminescence 14 days after inoculation of GL261-Luc into WT or p50−/− mice (mean and SE from four determinations). b Representative Foxp3;CD25 FC plots within the CD3+CD4+ T cell subset (left) and quantification of Foxp3+CD25+ Tregs as a percent of CD4 cells (right, mean and SE from four determinations) are shown. c Representative CD4;IFNγ and CD8;IFNγ FC plots, within the CD3+ T cell gate, are shown 4 hrs after exposure to vehicle or PMA/ionomycin (left). CD4+IFNγ+ or CD8+IFNγ+ cells were quantified as the relative number per bioluminescence unit (right, mean and SE from four determinations). d Representative CD3;IFNγ, CD3;Granzyme B (GrzB), and CD3;TNFα FC plots, within CD4+ or CD8+ gates, are shown three days after stimulation of isolated CD3+ tumor cells with Dynabeads containing CD3 and CD28 antibodies (left). IFNγ+, GrzB+, and TNFα+ cells were quantified as percent of CD4+ or CD8+ cells (right, mean and SE from five determinations).

Article Snippet: Cells were then either stained for flow cytometry (FC), or separated into CD1 lb + and CD1lb - or CD3 + and CD3 - cell fractions using CD1lb or CD3 positive selection kits and LS columns (Miltenyi).

Techniques: Isolation