anti vegfr2  (R&D Systems)

Journal: Frontiers in Cell and Developmental Biology

Article Title: Tissue-resident macrophages co-develop with myocardial tissue in human induced pluripotent stem cell-derived organoids

doi: 10.3389/fcell.2025.1629988

Figure Lengend Snippet: Cardiac Organoids. (A) Schematic representation of the cardiac organoid differentiation protocol displaying experimental timing, cell culture media as well as small molecules and cytokines used to induce cardiac tissue. The image was created in BioRender (Wörsdörfer, P. (2025) https://BioRender.com/rz8x29g ). (B) Semiquantitative RT-PCR detecting mesodermal ( TBXT , KDR , GATA4 , RUNX1 ), cardiac ( NKX2.5 , TBX5 , ISL1 , TNNT2 , DES , RYR2 ), and endothelial ( PECAM1 ) marker gene expression at days 2, 4 and 8 of differentiation. (C) H&E staining of an organoid section at day 2. (D,D′) Immunofluorescence analysis of the organoid shown in C detecting early mesodermal markers TBXT and VIM at day 2 of organoid development. (E) Immunofluorescence analysis of the organoid shown in C detecting early mesodermal markers GATA6 and KDR at day 2 of organoid development. (F) H&E staining showing hollow cavity-like structures in cardiac organoids at day 7. (G,G′) Immunofluorescence analysis of the organoid shown in F detecting mesenchymal cells (VIM + ) in close association with cardiomyocytes (TNNT2 + ) at day 7 of organoid development. (H) H&E staining of organoid sections at day 35 of development. (I) Immunofluorescence analysis of the organoid shown in H showing close interaction of fibroblasts (VIM + ) and cardiomyocytes (TNNT2 + ) at day 35 of organoid development. (J) Cardiac organoids progressively increase in size (day 0 = 712.2 ± 6.8 µm, n = 72; day 7 = 1,231 ± 20.3 µm, n = 70; day 25 = 1,613 ± 45.4 µm, n = 41. ***p < 0.001, ****p < 0.0001. (K) Contraction frequency of organoids increases over time, from 25 ± 1.9 bpm at day 7–57 ± 10.5 bpm at day 35 (n = 8). **p < 0.01, ****p < 0.0001.

Article Snippet: Primary antibodies against PECAM1 (CD31) (DAKO, M0823), VIM (Invitrogen, MA5-16409), ACTN1 (Abcam, Ab68167), KDR (Miltenyi Biotec, 130-125-988), TBXT (R&D Systems, AF2085), TNNT2 (Life Technologies, MA512960 ), CD34 (Life Technologies, MA5-32059), AIF1 (WAKO/Fuji Film, 019-19741) and PTPRC (CD45) (Life Technologies, 14-0451-82) were diluted in NBS blocking solution and incubated overnight at 4 °C.

Techniques: Cell Culture, Reverse Transcription Polymerase Chain Reaction, Marker, Gene Expression, Staining, Immunofluorescence

Journal: STAR Protocols

Article Title: Protocol for inducing beige adipocytes in white adipose tissue of mouse using cold exposure and CL316,243 injection

doi: 10.1016/j.xpro.2024.103337

Figure Lengend Snippet:

Article Snippet: Mouse VEGFR2/KDR/Flk-1 antibody (1: 200) , R&D Systems , Catalog # AF644, RRID: AB_355500.

Techniques: Recombinant, Avidin-Biotin Assay, Blocking Assay, Protease Inhibitor, Bicinchoninic Acid Protein Assay, Reverse Transcription, Software, Microscopy

Journal: Nature Cell Biology

Article Title: Specialized post-arterial capillaries facilitate adult bone remodelling

doi: 10.1038/s41556-024-01545-1

Figure Lengend Snippet: a , b , High-resolution confocal images showing EMCN + VEGFR3 − rECs (arrowheads) around TB in relation to RUNX2 + osteoprogenitors (green) ( a ) and ATP6V1B1/B2 + osteoclasts (green) ( b ). c , d , Maximum-intensity projection showing Sp7-mCherry + osteoblasts (yellow) and ATP6V1B1/B2 + osteoclasts (green) in in relation to EMCN + VEGFR3 − rECs around 6-week-old femoral TB ( c ). Single xy plane ( d ). Green arrowheads mark rECs near osteoclasts and yellow arrowheads near osteoblasts. e , f , Distribution of EMCN + VEGFR3 − type R vessels in relation to Sp7-mCherry + osteoblasts (yellow arrowheads) and ATP6V1B1/B2 + osteoclasts (green arrowheads) around 6-week-old and 12-week-old TB ( e ). Isolated xy plane ( f ). g , Confocal images showing VEGFR3 − VEGFR2 + ECs (white arrowheads) around TB in young and aged patient samples. VEGFR3 (yellow), VEGFR2 (red) and nuclei (DAPI, blue). DAPI, 4,6-diamidino-2-phenylindole.

Article Snippet: Tissues were incubated with primary antibodies rabbit anti-DACH1 (Proteintech, 10914-1-AP, 1:500 dilution), goat anti-VEGFR2 (R&D Systems, AF644; 1:100 dilution) followed by secondary antibodies anti-rabbit Alexa-488 (Thermo Fisher Scientific, A21206; 1:300 dilution) and anti-goat Alexa-594 (Thermo Fisher Scientific, A11058; 1:300 dilution) overnight on a rotating platform at 4 °C, incubated and mounted with Histodenz (Sigma, D2158) in DAPI.

Techniques: Isolation

Journal: Nature Cell Biology

Article Title: Specialized post-arterial capillaries facilitate adult bone remodelling

doi: 10.1038/s41556-024-01545-1

Figure Lengend Snippet: a, b . High-resolution confocal images showing VEGFR3 (yellow) and VEGFR2 (red) immunostaining together with DAPI (blue). VEGFR3 − VEGFR2 + capillaries (white arrowheads) around trabecular bone (TB) and nearby VEGFR3 + VEGFR2 + vessels (yellow arrowheads) in samples from young ( a ) and aged patients ( b ) are indicated. Scale bars, 200μm. c . Tile scan confocal images of EMCN + VEGFR3 − ECs in 12-week-old female and male murine femur. Trabecular bone (TB) is indicated. Scale bars, 500μm. d . Quantitation of EMCN + VEGFR3 − vessel density in 12-week-old female and male femur (n =3 mice per group). Mean ± SEM. P values, unpaired two-tailed t-test. e . EMCN (red) and VEGFR3 (blue) immunostaining showing the presence of VEGFR3 − EMCN + vessels (white arrowheads) around trabecular bone (TB) in female and male femur. Scale bars, 100μm.

Article Snippet: Tissues were incubated with primary antibodies rabbit anti-DACH1 (Proteintech, 10914-1-AP, 1:500 dilution), goat anti-VEGFR2 (R&D Systems, AF644; 1:100 dilution) followed by secondary antibodies anti-rabbit Alexa-488 (Thermo Fisher Scientific, A21206; 1:300 dilution) and anti-goat Alexa-594 (Thermo Fisher Scientific, A11058; 1:300 dilution) overnight on a rotating platform at 4 °C, incubated and mounted with Histodenz (Sigma, D2158) in DAPI.

Techniques: Immunostaining, Quantitation Assay, Two Tailed Test

Journal: Nature Cell Biology

Article Title: Specialized post-arterial capillaries facilitate adult bone remodelling

doi: 10.1038/s41556-024-01545-1

Figure Lengend Snippet: a , t -SNE visualization of Dach1 gene expression in ECs of different organs. b , Confocal tile scan overview images of 12-week-old Dach1 iΔEC and control retina stained for CD31 (red). Scale bars, 300μm. c , Quantitation of CD31 + vessel density in 12-week-old Dach1 iΔEC retinas relative to control (n =4 mice per group). Mean ± SEM. P values, unpaired two-tailed t-test. d , High-resolution confocal images of Dach1 iΔEC and control intestinal villi stained for DACH1 (green), VEGFR2 (red), and DAPI (blue). Scale bars, 50μm and insets 15μm. e , Quantitation of average VEGFR2 + vessel density, villus size, and average vascular area per villus in 12-week-old Dach1 iΔEC and littermate control (n =4 mice per group). Mean ± SEM. P values, unpaired two-tailed t-test. f , Confocal tile scan overview images of 12-week-old female Dach1 iΔEC and control femur stained for EMCN (red) and OSTERIX (green, OSX). White dashed lines and yellow arrowheads indicate reduction of region containing trabecular bone (TB) in Dach1 iΔEC mutants. Scale bars, 500μm. g . High-magnification images showing EMCN (red) and ATP6V1B1/B2 (green) immunostained female Dach1 iΔEC and control femur. Growth plate (GP) is indicated. Scale bars, 100μm. h . Quantitation of OSX + cells and ATPV6B1B2 area in control and Dach1 iΔEC mutant (n=3 and 4 mice per group). Mean ± SEM. P values, unpaired two-tailed t-test. i . Quantitation shows connectivity density and cortical thickness (per mm), (n = 3-4 female mice per group and n=3 male mice per group). Mean ± SEM. P values, plotted using unpaired two-tailed t-test. j . DACH1 (green) co-stained with VE-CADHERIN (red) in siCONTROL and siDACH1 cultured HUVECs. k . Downregulation of arterial marker genes in siDACH1 HUVECs relative to siCONTROL . n=6 (3 independent experiments) for each group. Mean ± SEM. P values, unpaired two-tailed t-test with Welch’s correction.

Article Snippet: Tissues were incubated with primary antibodies rabbit anti-DACH1 (Proteintech, 10914-1-AP, 1:500 dilution), goat anti-VEGFR2 (R&D Systems, AF644; 1:100 dilution) followed by secondary antibodies anti-rabbit Alexa-488 (Thermo Fisher Scientific, A21206; 1:300 dilution) and anti-goat Alexa-594 (Thermo Fisher Scientific, A11058; 1:300 dilution) overnight on a rotating platform at 4 °C, incubated and mounted with Histodenz (Sigma, D2158) in DAPI.

Techniques: Expressing, Control, Staining, Quantitation Assay, Two Tailed Test, Mutagenesis, Cell Culture, Marker