Journal:

Article Title: Expansion of activated human na?ve T-cells precedes effector function

doi: 10.1046/j.1365-2249.2002.02015.x

Figure Lengend Snippet: Surface markers and cytokines expressed by naïve and memory T-cells

Article Snippet: Antigen-experienced cells were purified by incubating cells with a FITC-conjugated antibody to CD45RA, then depleted by MACS sorting using anti-FITC magnetic microbeads (Miltenyi, Auburn, CA, USA).

Techniques: Expressing

Journal:

Article Title: Expansion of activated human na?ve T-cells precedes effector function

doi: 10.1046/j.1365-2249.2002.02015.x

Figure Lengend Snippet: .Stimulated antigen-experienced T-cells gain effector function without cell division. PBMC from healthy adult donors depleted of CD45RA+ T-cells, were stained with 0·25 µm CFSE and were then stimulated for 4 days with PHA or SEB, or for 6 days by MLR. Shown is a dot plot panel of surface markers and cytokine production for antigen-experienced CD4+ or CD8+ gated T-cells.

Article Snippet: Antigen-experienced cells were purified by incubating cells with a FITC-conjugated antibody to CD45RA, then depleted by MACS sorting using anti-FITC magnetic microbeads (Miltenyi, Auburn, CA, USA).

Techniques: Staining

Journal:

Article Title: Expansion of activated human na?ve T-cells precedes effector function

doi: 10.1046/j.1365-2249.2002.02015.x

Figure Lengend Snippet: IL-7 drives proliferation of naïve T-cells in MHC independent fashion. Cord blood PBMC were stained with 0·25 µm CFSE and were then cultured for 9 days in 10 ng/ml IL-7 with isotypic control antibodies (a) or 10 µg/ml blocking antibodies recognizing HLA A, B, C, DR, DP, and DQ (b) Dot plots of CD4+ or CD8+ gated T-cells show expression patterns of CD45RA, and CD45RO with respect to cell division. Boxed areas of CD4 and CD8 plots represent the percentage of cultured cells that had proliferated more than three times for IL-7 alone (a) and IL-7+ anti-MHC antibodies (b).

Article Snippet: Antigen-experienced cells were purified by incubating cells with a FITC-conjugated antibody to CD45RA, then depleted by MACS sorting using anti-FITC magnetic microbeads (Miltenyi, Auburn, CA, USA).

Techniques: Staining, Cell Culture, Control, Blocking Assay, Expressing

Journal:

Article Title: Expansion of activated human na?ve T-cells precedes effector function

doi: 10.1046/j.1365-2249.2002.02015.x

Figure Lengend Snippet: IL-2 is Required for IL-7 induced expansion of memory T-Cells present in umbilical cord blood. Cord blood PBMC were stained with 0·25 µm CFSE and were then cultured for 9 days in 10 ng/ml IL-7 with or without 10 µg/ml depleting antibodies to IL-2. After culturing, PBMC were stained with CD4 and CD45RO or CD45RA, and CD8 and CD45RA or CD45RO. Dot plots of CD4+ or CD8+ T-cells show expression patterns of CD45RA, CD45RO, CD25, and IFN-γ with respect to cell division.

Article Snippet: Antigen-experienced cells were purified by incubating cells with a FITC-conjugated antibody to CD45RA, then depleted by MACS sorting using anti-FITC magnetic microbeads (Miltenyi, Auburn, CA, USA).

Techniques: Staining, Cell Culture, Expressing

Journal: medRxiv

Article Title: Systemic blockade of Clever-1 elicits lymphocyte activation alongside checkpoint molecule downregulation in patients with solid tumors

doi: 10.1101/2020.11.11.20227777

Figure Lengend Snippet: A , T-distributed Stochastic Neighbor Embedding (tSNE) of CD3-excluded circulating cells from pre-dose (D0, n=7) and day-7 (D7, n=7) samples pre-gated for viability, singlets and CD45 + . B , tSNE heatmaps showing expression of indicated markers on cell clusters of a representative patient. C , Median expression of CD206 and CD163 on CD14 + monocytes at pre-dose and 7 days after FP-1305 treatment. Paired student’s t -test. D , Representative flow cytometry plots showing size (FSC) and granularity (SSC) of CD14 high monocytes (black), and median expression of CD14 and Clever-1 (9-11 ab) at different time points during treatment cycle 1. Clever-1 R-O signal was detected by alexa647 conjugated FP-1305 for investigating receptor (Clever-1) occupancy. One-way ANOVA performed between timepoints day 0, 1 and 7. E , Unsupervised hierarchical clustering of n=13,589 genes expressed in CD14 + monocytes obtained from four patients across timepoints day 0, 1 and 7. F , Principal component analysis of patient samples across different timepoints. G , Venn diagrams showing differentially expressed genes in comparison to pre-dose (D0). H , Ingenuity Pathway Analysis of patient gene expression changes at day 7 compared to pre-dose. Red color indicates predicted pathway activation and blue color inhibition. I , FPKM and RQ values of significantly altered genes at different timepoints. Friedman test. All P *<0.05.

Article Snippet: Thereafter the cells were barcoded with heavy-metal isotope-labelled anti-human CD45 (clone H130) antibodies (CD45_89Y (Fluidigm), CD45_141Pr (Fluidigm) and CD45_147Sm (BioLegend), 1/200) for 30 min at RT, washed carefully, combined and divided equally into two samples.

Techniques: Expressing, Flow Cytometry, Activation Assay, Inhibition