tryple express enzyme 12605010  (Thermo Fisher)

Journal: eLife

Article Title: GATA6 regulates WNT and BMP programs to pattern precardiac mesoderm during the earliest stages of human cardiogenesis

doi: 10.7554/eLife.100797

Figure Lengend Snippet:

Article Snippet: Other , TrypLE Express Enzyme , Thermo Fisher , 12605010 , Reagent.

Techniques: Binding Assay, Mutagenesis, Transfection, Construct, CRISPR, Plasmid Preparation, Derivative Assay, Expressing, Control, Sequencing, Recombinant, T7EI Assay, DNA Extraction, DNA Purification, cDNA Synthesis, SYBR Green Assay, Illumina Sequencing, Multiplex Assay, Bicinchoninic Acid Protein Assay, Western Blot, Selection, Software, Knock-Out, Lysis, Electron Microscopy

Journal: eLife

Article Title: Evaluation of Gremlin-1 as a therapeutic target in metabolic dysfunction-associated steatohepatitis

doi: 10.7554/eLife.95185

Figure Lengend Snippet: ( A ) Representative RNAscope in situ hybridisation (ISH) images for co-staining of GREM1 (red) and THY1 (green) in normal human liver and MASH fibrosis. Scale bar represents 100 μM. ( B ) Quantification of ISH staining areas across different stages of liver fibrosis. Significance was assessed by two-sided Jonckheere-Terpstra test (***p=1.3 × 10 –09 ). ( C ) Quantification of human GREM1 qPCR across chronic liver diseases of different aetiology. Data are given as mean -ΔΔCt ± SD, relative to donor liver and normalised to the expression of SRSF4, HPRT1, and ERCC3. Significance was assessed by multiple two-sided Welch’s t-test against donor control, followed by Bonferroni-Holm adjustment (*p<0.05, **p=0.004). ( D ) Representative histological images of RNAscope in situ hybridisation (ISH) for co-staining of GREM1 (red) and THY1 or COL3A1 (green) in MASH fibrosis. Representative double positive cells are indicated by arrows. Scale bar represents 50 μM. ( E ) Quantification of qPCR for GREM1 mRNA in major primary human non-parenchymal cell types. HSEC – human sinusoidal endothelial cells, BEC – biliary epithelial cells, HHSC – human hepatic stellate cells, MF – myofibroblasts. ( F ) Representative RNAscope ISH images for GREM1 (red) in rats fed a standard chow or choline-deficient, L-amino acid defined high-fat diet (CDAA-HFD) for 12 weeks. Scale bar represents 50 μM. Figure 1—source data 1. Excel spreadsheet containing data displayed in panels A, C, and E.

Article Snippet: Myofibroblasts were grown on uncovered polystyrene culture plates in 16% FCS in DMEM supplemented with 1% penicillin-streptomycin-L-glutamine and subcultured at a 1:3 ratio using Gibco TrypLE Express Enzyme (12605010, Fisher Scientific) for cell dissociation.

Techniques: RNAscope, In Situ, Hybridization, Staining, Expressing, Control

Journal: eLife

Article Title: Evaluation of Gremlin-1 as a therapeutic target in metabolic dysfunction-associated steatohepatitis

doi: 10.7554/eLife.95185

Figure Lengend Snippet: ( A ) Fibrogenic marker genes in primary human hepatic stellate cells treated with anti-Gremlin-1 (aG1) or isotype control antibodies (iso-Ab). ( B ) Fibrogenic marker genes in primary human hepatic myofibroblasts treated with anti-Gremlin-1 or isotype control antibodies. ( C ) Fibrogenic gene expression in lentivirally transduced human hepatic stellate cells (HHSC). ( D ) Fibrogenic gene expression in lentivirally transduced LX-2. ( E ) Bone morphogenetic protein (BMP) signalling-related gene expression in lentivirally transduced LX-2. ( A–B ) Data are presented as individual data points and mean for -ΔΔCt relative to untreated control and normalised to the expression of SRSF4. *p<0.05 in one-way ANOVA and post hoc paired t-tests for pre-defined comparisons with Bonferroni-Holm adjustment. ( C–E ) Data are given as mean ± SEM of -ΔΔCt relative to GFP and vehicle control and normalised to the expression of SRSF4. *p<0.05 in GREM1 vs GFP-control, #p<0.05 in TGFβ1 vs vehicle control in repeated measures two-way ANOVA and post hoc paired t-test for pre-selected comparisons and Bonferroni-Holm adjustment. Figure 4—source data 1. Excel spreadsheet containing data displayed in panels A–E.

Article Snippet: Myofibroblasts were grown on uncovered polystyrene culture plates in 16% FCS in DMEM supplemented with 1% penicillin-streptomycin-L-glutamine and subcultured at a 1:3 ratio using Gibco TrypLE Express Enzyme (12605010, Fisher Scientific) for cell dissociation.

Techniques: Marker, Control, Gene Expression, Expressing