Journal: Development (Cambridge, England)

Article Title: Understanding human fetal pancreas development using subpopulation sorting, RNA sequencing and single-cell profiling

doi: 10.1242/dev.165480

Figure Lengend Snippet: Characterization of sorted cell populations from human fetal pancreata by RNA-seq at 9 WD. (A) Flow cytometry analysis of the expression of GP2 and ECAD on CD45−CD31−EPCAM+ cells at 9 WD. CD142 and SUSD2 expression was analyzed in GP2−ECAD+ and GP2−ECAD− cells. FACS plots are representative of three independent pancreata at 9 WD. (B) Scheme of the experimental setup: GP2−ECAD+CD142+SUSD2− (population A, yellow), GP2−ECAD+CD142−SUSD2− (population B, orange), GP2−ECADlowCD142−SUSD2+ (population C, red) and GP2−ECADlowCD142−SUSD2− (population D, blue) were cell sorted and analyzed by RNA-seq. All populations were CD45−CD31−EPCAM+ and derived from three independent pancreata at 9 WD. The population color code is re-used throughout the article. (C) Principal component analysis map on the RNA-seq from all four sorted populations in triplicate at 9 WD. (D) Heatmap displaying the expression of the 1007 differentially expressed genes in triplicate in all four populations in RPKM, along with the hierarchical clustering. Clusters are named on the right side and some gene examples are indicated in boxes. For more extensive examples, Table S1 highlights genes enriched in the four sorted cell populations.

Article Snippet: The following antibodies were used: anti-CD45-PerCP/Cy5.5 (1/20, clone 2D1, BioLegend), anti-CD31-PerCP/Cy5.5 (1/20, clone WM59, BioLegend), anti-CD235a-PerCP/Cy5.5 (1/20, clone HI264, BioLegend), anti-EPCAM-Brillant Violet 605 (1/20, clone 9C4, BioLegend), anti-ECAD-PE-Cy7 (1/20, clone 67A4, BioLegend), anti-CD133-APC (1/20, clone 315-2C11, BioLegend), anti-GP2-PE (1/5, clone 3G7H9, MBL International), anti-SUSD2-VioBrightFITC (1/20, W5C5, Miltenyi Biotec), anti-CD142-BV711 (1/20, clone HTF-1, BD Biosciences), anti-NKX6-1-PE (1/40, clone R11-560, BD Biosciences), sheep anti-NEUROG3 (1/300, AF3444, R&D Systems), mouse anti-NKX2-2 (1/200, F4.5A5, Developmental Studies Hybridoma Bank), C-peptide-Alexa Fluor 647 (1/200, clone U8-424, BD Biosciences) and Glucagon-BV421 (1/80, clone U16-850, BD Biosciences).

Techniques: RNA Sequencing Assay, Flow Cytometry, Expressing, Derivative Assay

Journal: American Journal of Transplantation

Article Title: Characterization of ASKP1240, a Fully Human Antibody Targeting Human CD40 With Potent Immunosuppressive Effects

doi: 10.1111/ajt.12678

Figure Lengend Snippet: Influence of ASKP1240 on human platelet and endothelial cell activation markers. (A) Effect of ASKP1240 on platelet P-selectin (CD62P) expression. Diluted PRP was incubated with indicated reagents. Shown are dot plots obtained from flow cytometory analyses demonstrating the cells incubated only media, 100 µM of thrombin receptor activating peptide (SFLLRN), 1 mg/mL of ASKP1240, 1 µg/mL of shCD154 or 1 mg/mL of ASKP1240 plus 1 µg/mL of shCD40. (B) Effect of ASKP1240 on endothelial cell E-selectin (CD62E) and tissue factor (CD142) expression. Shown are dot plots obtained from flow cytometory analyses demonstrating the cells incubated with only media, 100 U/mL of TNFα, 1 mg/mL of ASKP1240, 1 µg/mL of shCD154 or 1 mg/mL of ASKP1240 plus 1 µg/mL of shCD40. PRP, platelet-rich plasma; shCD154, soluble human CD154; TNF, tumor necrosis factor.

Article Snippet: Following incubation, the cells were collected by trypsinization, stained with PE-labeled anti-CD62E (clone 68-5H11; BD) or PE-labeled anti-CD142 (clone HTF-1; BD) and fluorescence activated cell sorting analyses were performed.

Techniques: Activation Assay, Expressing, Incubation