Paraffin- or optimal cutting temperature compound-embedded tissues were sectioned and stained6 (link)7 (link) using the following primary antibodies (all diluted 1:100 unless stated otherwise) for immunofluorescence labelling: Lrig: R&D Systems, FAB3688G; CD26: R&D Systems, AF954; Sca1: R&D Systems, AF1226; PDGFRa: R&D Systems, AF1062; Collagen III: Abcam, ab7778, Collagen11a1: Abcam, ab64883; Elastin: Abcam, ab21610; Caveolin: Cell Signaling Technology, 3267; phospho-Histone H3 (Ser10) antibody: Cell Signalling Technology, 970; Active Caspase-3: RnD Systems, AF835; K14: Covance, PRB-155P, 1:500; GFP: Abcam, ab13970, 1:500; RFP: Rockland, 600-401-379, 1:300. EdU staining was performed with a Click-it EdU imaging kit (Invitrogen) according to the manufacturer's recommendations. Images were acquired with a Nikon A1 Upright Confocal microscope. Images of H&E- and Herovici-stained sections were acquired with a Hamamatsu slide scanner. Representative images of skin from two to three independent experiments with at least three biological replicates per group are shown.
Pdgfra
Manufactured by R&D Systems
Sourced in Canada
PDGFRa is a receptor tyrosine kinase that binds platelet-derived growth factor (PDGF). It plays a role in cell proliferation, survival, and migration.
Automatically generated - may contain errors
Lab products found in correlation
Ab64883, by Abcam (1 mentions)
Slide scanner, by Hamamatsu Photonics (1 mentions)
Ab13970, by Rockland Immunochemicals (1 mentions)
Ab7778, by Abcam (1 mentions)
Elastin, by Abcam (1 mentions)
Collagen 3, by Abcam (1 mentions)
A1 upright confocal microscope, by Nikon (1 mentions)
Click it edu imaging kit, by Thermo Fisher Scientific (1 mentions)
Fab3688g, by R&D Systems (1 mentions)
Af954, by R&D Systems (1 mentions)
Sca 1, by R&D Systems (1 mentions)
Af1226, by R&D Systems (1 mentions)
Prb 155p, by Abcam (1 mentions)
Af1062, by Abcam (1 mentions)
Phospho histone h3 ser10 antibody, by Cell Signaling Technology (1 mentions)
Perilipin, by Abcam (1 mentions)
Active caspase 3, by Cell Signaling Technology (1 mentions)
Prolong gold with dapi, by Thermo Fisher Scientific (1 mentions)
Lsm 710 meta, by Zeiss (1 mentions)
Alexa fluor 488 donkey anti mouse, by Thermo Fisher Scientific (1 mentions)
5 protocols using pdgfra
1
Immunostaining of Skin Sections
2
Immunohistochemical Analysis of Embryonic and Skin Samples
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Prefixed (4% in PFA) OCT-embedded embryos or skin samples were sectioned at the desired thickness (10–40 µm). Immunohistochemistry was performed on sectioned slides by primary antibody incubation overnight at 4°C followed by secondary antibody staining for 1–4 h at room temperature. The following antibodies and dilutions were used: active-Caspase-3 (rabbit; 1:600; Cell Signaling Technology), GFP (rabbit; 1:400; Abcam), Pdgfra (goat; 1:200; R&D), Dlk (goat; 1:200; R&D), Perilipin (goat; 1:800; Abcam), CD31 (rat; 1:100; eBioscience), and CD45 (rat; 1:100; eBioscience). The secondary antibodies used were donkey anti-goat, rabbit, or rat conjugated with Alexa 488, 549, or 647 (1:800; Jackson Immunoresearch), respectively. EdU Click-It reaction was performed for 1 h at room temperature according to the manufacturer's instructions (Life Technologies). Samples were mounted in Prolong Gold with DAPI (Life Technologies).
3
Histological Analysis of Mouse Skin
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Dorsal skin of mice was processed for histological sections, and 5-μm-thick paraffin sections were subjected to H/E staining or immunohistochemical analysis. The antibodies used for immunochemical detection were anti-15-lipoxygenase-1 antibody (Abcam), Perilipin A (Cell Signaling), Keratin 14 (Cell Signaling), Keratin 15 (Cell Signaling), PDGFRA (R&D systems), smooth muscle actin-FICT (Sigma), HMGB1 (Cell Signaling), tdTomato (Clonetech), Ly6C (Biolegend) and F4/80 antibody (AbD Serotec). The secondary antibodies used were donkey anti-mouse-Alexa Fluor 488, donkey anti-rabbit-Alexa Fluor 594/Alexa Fluor 488, donkey anti-goat-Alexa Fluor 594, and donkey anti-rat-Alexa Fluor 594 (ThermoFisher Scientific, Molecular Probes). The omission of primary antibody or normal rabbit, rat, goat, or mouse IgG controls (Santa Cruz Biotechnology) was used as a negative control. DAPI (Sigma) was used for nuclear counterstaining. Cells were imaged on a Zeiss confocal laser-scanning microscope (LSM 710 META, Zeiss, Jena, Germany).
4
Comprehensive Protein Analysis of Cellular Lysis
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Cells were lysed in RIPA lysis buffer supplemented with phosphatase inhibitor tablets (Roche, 04,906,837,001) and protease inhibitor tablet (Roche, 05,892,970,001). Primary antibodies were Vimentin (Cell Signaling, 5741,1:1000), DDR2 (R&D, MAB25381, 1:2000), PDGFRA (R&D, AF1062, 1:1000), Periostin (R&D, AF2955, 1:1000), Wnt5a (Cell Signaling, 2530, 1:1000), β-Actin (Cell Signaling, 4970, 1:2000), β-Catenin (Cell Signaling, 8480, 1:2000), α-SMA (Sigma, A5228, 1:2000), Fibronectin (Cell Signaling, 26,836, 1:1000), Collagen I (abcam, ab6308, 1:1000), GAPDH (Cell Signaling, 5174, 1:2000), Smad2 (Cell Signaling, 5339, 1:1000), Phospho- Smad2 (Cell Signaling, 3108, 1:1000).
5
Immunohistochemical Profiling of Organoid Structures
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Four-micron sections of formalin-fixed paraffin embedded organoids were stained with hematoxylin and eosin (Ventana Medical Systems, Tucson, AZ). Immunohistochemistry was performed with an automated, validated, and accredited staining system (Ventana Benchmark ULTRA) using the UltraView or OptiView universal DAB detection Kit (Ventana Medical Systems). After deparaffinization and heat-induced antigen retrieval, the tissue samples were incubated with WT1 (Cell Marque, Rocklin, CA), ECAD (Ventana Medical Systems), Villin-1 (Abcam, Cambridge, UK), CD31 (Cell Marque), renin (Abcam), NKCC2 (StressMarq Biosciences, Victoria, BC, Canada), NHE3 (StressMarq), CD34 (Cell Marque), COL1A1 (Novus Biologicals, Littleton, CO), or PDGFRa (R&D systems). Incubation was followed by hematoxylin II counterstaining. Double stainings of PDGFRa and renin were performed by incubating the samples with PDGFRa for 32 minutes at 37 C followed by detection with Red610 kit (#760-245, Ventana) and renin for 32 minutes at 37 C followed by detection with FAM kit (#760-243, Ventana). An AF488-conjugated anti-human mitochondria antibody MAB1273A4 (Millipore, Billerica, MA) was used to detect
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