pdx 1  (Proteintech)

Journal: bioRxiv

Article Title: Iterative sacrificial 3D printing and polymer casting to create complex vascular grafts and multi-compartment bioartificial organs

doi: 10.1101/2024.09.29.615298

Figure Lengend Snippet: a , Illustration of the protocol used to differentiate stem cell towards SC-islets. Illustration made with Bio-render. b , Representative brightfield images showing SC-islets after 10 days of culture (Stage 7) inside the device either in static or perfused condition. Scale bars, 200 µm. c , Representative fluorescent images of live/dead staining of the pseudoislets after perfusion, with Calcein AM (green) and propidium iodide (red). Scale bars, 200 µm. d , Hematoxylin and Eosin staining of the SC-islets after 10 days of perfusion. Scale bars, 200 µm. e , Representative dithizone staining (crimson red) of the SC-islets after 10 days of culture. Scale bars, 200 µm. f , Representative fluorescent images of SC-islets after maturation in the device. Cells are labelled with dapi (blue), NKX6.1 or PDX1 (green) and insulin or glucagon (pink). Scale bars, 100 µm. g , Flow cytometry results showing the percentage of the population expressing key pancreatic and endocrine markers. Mean and standard deviation are shown for 3 independent devices with three independent differentiations. *p < 0.05, determined by two-way ANOVA with Tukey’s multiple comparisons test. h , Gene expression of the SC-islets after 10 days of perfusion. Mean and standard deviation are shown for 3 independent devices with three independent differentiations. n=3 technical replicates per point. ns=non-significant, determined by a two-way ANOVA with Šídák’s multiple comparisons test. i , Insulin secretion in response to changes in glucose concentration or addition of KCl, normalized to the total insulin content. Mean and standard deviation for three independent differentiations with 3 independent devices are shown.

Article Snippet: Cells were then incubated with conjugated antibodies for 30 minutes: SOX17-PE (561591, BD Biosciences), PDX1-PE (562161, BD Biosciences), NKX6.1-AF647 (563338, BD Biosciences), NEUROD1-PE (563001, BD Biosciences), glucagon-PE (565860, BD Biosciences), c-peptide-AF647 (565831, BD Biosciences), AlexaFluor® 647 mouse IgG1 K Isotype Control (557732, BD Biosciences), PE mouse IgG1 K Isotype Control (554680, BD Biosciences).

Techniques: Staining, Flow Cytometry, Expressing, Standard Deviation, Concentration Assay

Journal: bioRxiv

Article Title: Iterative sacrificial 3D printing and polymer casting to create complex vascular grafts and multi-compartment bioartificial organs

doi: 10.1101/2024.09.29.615298

Figure Lengend Snippet: a , Illustration of the protocol used to differentiate stem cell towards SC-islets. Illustration made with Bio-render. b , Representative brightfield images showing SC-islets after 10 days of culture (Stage 7) inside the device either in static or perfused condition. Scale bars, 200 µm. c , Representative fluorescent images of live/dead staining of the pseudoislets after perfusion, with Calcein AM (green) and propidium iodide (red). Scale bars, 200 µm. d , Hematoxylin and Eosin staining of the SC-islets after 10 days of perfusion. Scale bars, 200 µm. e , Representative dithizone staining (crimson red) of the SC-islets after 10 days of culture. Scale bars, 200 µm. f , Representative fluorescent images of SC-islets after maturation in the device. Cells are labelled with dapi (blue), NKX6.1 or PDX1 (green) and insulin or glucagon (pink). Scale bars, 100 µm. g , Flow cytometry results showing the percentage of the population expressing key pancreatic and endocrine markers. Mean and standard deviation are shown for 3 independent devices with three independent differentiations. *p < 0.05, determined by two-way ANOVA with Tukey’s multiple comparisons test. h , Gene expression of the SC-islets after 10 days of perfusion. Mean and standard deviation are shown for 3 independent devices with three independent differentiations. n=3 technical replicates per point. ns=non-significant, determined by a two-way ANOVA with Šídák’s multiple comparisons test. i , Insulin secretion in response to changes in glucose concentration or addition of KCl, normalized to the total insulin content. Mean and standard deviation for three independent differentiations with 3 independent devices are shown.

Article Snippet: The samples were then incubated overnight in humidified chamber at 4 °C in the presence of antibodies against VE-cadherin (1:200, R&D, catalog no. MAB9381), NKX6.1 (1:100, DSHB, catalog no. F55A10), PDX1 (1:200, R&D, catalog no.MAB2419), insulin (1:200, Invitrogen, catalog no. 701265) or glucagon (1:200, Invitrogen, catalog no. 14-9743-82) in antibody diluent (Dako).

Techniques: Staining, Flow Cytometry, Expressing, Standard Deviation, Concentration Assay