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AF762

Tie-2 Antibody [Unconjugated]

(2 Reviews)

(36 Publications)

100 μg

AF762

Tie‑2 was detected in immersion fixed paraffin-embedded sections of human intestine tissue using Goat Anti-Mouse/Rat Tie‑2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF762) at 1 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cell surface on endothelial cells. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Pericytes express functional Tie2.(a) Microarray-based expression screening of brain pericytes (BP), placenta pericytes (PP), pancreas pericytes (PA), lung pericytes (LP), muscle pericytes (MP) and human umbilical vein endothelial cells (HUVEC [HU]). (b) Semi-quantitative PCR of TEK (Tie2), endothelial marker genes (PECAM1, CDH5) and pericyte marker genes (CSPG4, CD248) in HUVEC and BP; house keeping gene: GAPDH. (c) Representative images showing expression of Tie2 in HUVEC and BP. (d) Histogram of membrane-bound Tie2 expression in HUVEC and BP compared to isotype control measured by flow cytometry. (e,g) Western blot (WB) analysis of tyrosine phosphorylation (pTyr) and total Tie2 after Tie2 immunoprecipitation (IP) in BP (e) and HUVEC (g) upon stimulation with recombinant human (rh) Ang1 compared to unstimulated (us) cells. (f,h) Quantification of the ratio of phosphorylated Tie2 (pTie2) relative to total Tie2 protein in BP (f) and HUVEC (h) normalized to us control (n=3). (i,k) Western blot analysis of AKT phosphorylation (Ser473) in control (shCtr) and Tie2-silenced (shTie2 I/shTie2 II) BP (i) and HUVEC (k) upon stimulation with rhAng1 or rhAng2. (j,l) Quantification of the ratio of pAKT to total AKT protein expression in BP (j) and HUVEC (l) normalized to unstimulated control (n=3). Representative western blot images are cropped versions and original images can be found in Supplementary Fig. 15. Scale bars: 20 μm (c). Data are shown as mean±s.d. Statistics were performed using Mann–Whitney U test (f,h) and one-way ANOVA (j,l). *P<0.05, **P<0.01. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/28719590), licensed under a CC-BY license. Not internally tested by R&D Systems.

Endothelial changes after pericyte depletion. a–f Maximum intensity projection of confocal images from control and DTRiPC P6 retinas stained for IB4 (red) in combination with VEGF-A a, VEGFR2 b, VEGFR3 c, Tie2 d, Esm1 e, and Dll4 f (all in white), as indicated. Note local increase of VEGFR2, VEGFR3, and Esm1 (arrowheads in b, c, e) but not Tie2 or VEGF-A at the edge of the vessel plexus. Dll4 expression in DTRiPC sprouts is increased in some regions (arrowheads) but absent in others (arrows). Scale bar, 100 µm. g–j Quantitation of VEGF-A immunosignals area and intensity g, signal intensity for VEGFR2 h and VEGFR3 i and proportion of Esm1+ area with respect to vascular area j in the P6 control and DTRiPC angiogenic front. Error bars, s.e.m. p-values, Student’s t-test. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/29146905), licensed under a CC-BY license. Not internally tested by R&D Systems.

Pericyte-derived Angpt1 controls alveologenesis. a RT-qPCR analysis of Angpt1 and Tie2/Tek expression in freshly sorted lung GFP+, CD31+ or EpCAM+ cells from P7 Pdgfrb(BAC)-CreERT2 R26-mT/mG mice. Data represents mean ± s.e.m. (n = 4 mice). b High magnification images of P10 Angpt1GFP lungs stained for GFP (green), PDGFR beta (red), and PDGFR alpha (blue). Arrows indicate GFP and PDGFR beta double positive pericytes. Scale bar, 15 µm. c RT-qPCR analysis of Angpt1 expression in freshly sorted PDGFR beta + cells from P7 Yap1,Wwtr1iPCKO and control lungs. Data represents mean ± s.e.m. (n = 4 mice, two-tailed unpaired t-test). dAngpt1 expression in cultured Verteporfin (VP)-treated (48 h) and control pericytes. Data represents mean ± s.e.m. (n = 4, Welch’s t-test). e Expression of the indicated transcripts in freshly sorted CD31+ cells from P7 Yap1,Wwtr1iPCKO and control lungs. Data represents mean ± s.e.m. (n = 4 mice, NS not significant, two-tailed unpaired t-test). f–h Western blot analysis of Angpt1 protein (f; n = 2 controls and 4 mutant mice) and of total and phospho-Tie2 (pTie2) in P12 Yap1,Wwtr1iPCKO and control total lung lysates (g, n = 3 controls and 5 mutants). Molecular weight marker (kDa) is indicated. Relative quantification of signals is shown in h. Two-tailed unpaired t-test. i Scheme showing the time points of tamoxifen administration and analysis for Angpt1iPCKO mice. j, k 3D reconstruction confocal images of P12 Angpt1iPCKO and littermate control lungs stained for AQP5 (green), PDGFR beta (red), and PECAM1 (blue). Panels in k show higher magnification of PECAM1 staining. Scale bar, 50 µm (j) and 30 µm (k). l Quantitation of airspace volume in P12 Angpt1iPCKO and littermate control lung sections with 3D reconstruction surface images. Data represents mean ± s.e.m. (n = 4 mice; p < 0.0001, two-tailed unpaired t-test). m 3D reconstruction confocal images of P12 Angpt1iPCKO and littermate control lungs stained for alpha SMA (red) and tropoelastin (blue). Scale bar, 50 µm. n Quantitation of staining intensity for alpha SMA or tropoelastin shown in m. Intensity was normalized to the size of the parenchymal region. Data represents mean ± s.e.m. (n = 4 mice; NS not significant, two-tailed unpaired t-test). o Schematic summary of findings. Pulmonary pericytes regulate ECs and alveolar epithelial cells via angiocrine factors such as angiopoietin-1 and HGF. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/29934496), licensed under a CC-BY license. Not internally tested by R&D Systems.

Pericytes express functional Tie2.(a) Microarray-based expression screening of brain pericytes (BP), placenta pericytes (PP), pancreas pericytes (PA), lung pericytes (LP), muscle pericytes (MP) and human umbilical vein endothelial cells (HUVEC [HU]). (b) Semi-quantitative PCR of TEK (Tie2), endothelial marker genes (PECAM1, CDH5) and pericyte marker genes (CSPG4, CD248) in HUVEC and BP; house keeping gene: GAPDH. (c) Representative images showing expression of Tie2 in HUVEC and BP. (d) Histogram of membrane-bound Tie2 expression in HUVEC and BP compared to isotype control measured by flow cytometry. (e,g) Western blot (WB) analysis of tyrosine phosphorylation (pTyr) and total Tie2 after Tie2 immunoprecipitation (IP) in BP (e) and HUVEC (g) upon stimulation with recombinant human (rh) Ang1 compared to unstimulated (us) cells. (f,h) Quantification of the ratio of phosphorylated Tie2 (pTie2) relative to total Tie2 protein in BP (f) and HUVEC (h) normalized to us control (n=3). (i,k) Western blot analysis of AKT phosphorylation (Ser473) in control (shCtr) and Tie2-silenced (shTie2 I/shTie2 II) BP (i) and HUVEC (k) upon stimulation with rhAng1 or rhAng2. (j,l) Quantification of the ratio of pAKT to total AKT protein expression in BP (j) and HUVEC (l) normalized to unstimulated control (n=3). Representative western blot images are cropped versions and original images can be found in Supplementary Fig. 15. Scale bars: 20 μm (c). Data are shown as mean±s.d. Statistics were performed using Mann–Whitney U test (f,h) and one-way ANOVA (j,l). *P<0.05, **P<0.01. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/28719590), licensed under a CC-BY license. Not internally tested by R&D Systems.

Reactivity:

Mu, Rt

Species Glossary

Reactivity: Mu, Rt

Applications:

,

Applications: ,

Host:

Goat Polyclonal

Host: Goat Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

Available Conjugates:

Conjugate	Catalog #	Availability	Size	Price
Biotin	BAF762

Available Formulations

Formulation	Catalog #	Availability	Price
Carrier Free	AF762-SP

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AF2720

Tie-2 [p Tyr992] Antibody [Unconjugated]

(1 Review)

(32 Publications)

50 μg

AF2720

Western blot shows lysates of NIH-3T3 mouse embryonic fibroblast cell line transfected with mouse Tie-2 and untreated (-) or treated (+) with 600 ng/mL Recombinant Human Angiopoietin-1 (Catalog # 923-AN) for 5 minutes. PVDF membrane was probed with 0.5 µg/mL of Rabbit Anti-Human/Mouse Phospho-Tie-2 (Y992) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2720) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band was detected for Phospho-Tie-2 (Y992) at approximately 150 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Tie‑2 was detected in immersion fixed paraffin-embedded sections of human kidney tissue using Rabbit Anti-Human/Mouse Phospho-Tie‑2 (Y992) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF2720) at 1 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Rabbit IgG VisUCyte™ HRP Polymer Antibody (brown; Catalog # VC003) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm in convoluted tubules. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

HUVEC human umbilical vein endothelial cells were unstimulated (light orange filled histogram) or treated with 100 μM pervanadate for 15 minutes (dark orange filled histogram), then stained with Human/Mouse Phospho-Tie‑2 (Y992) Antigen Affinity‑purified Polyclonal Antibody (Catalog # AF2720) or control antibody (Catalog # AB-105-C, open histogram), followed by Phycoerythrin-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # F0110). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with methanol.

Pericyte-derived Angpt1 controls alveologenesis. a RT-qPCR analysis of Angpt1 and Tie2/Tek expression in freshly sorted lung GFP+, CD31+ or EpCAM+ cells from P7 Pdgfrb(BAC)-CreERT2 R26-mT/mG mice. Data represents mean ± s.e.m. (n = 4 mice). b High magnification images of P10 Angpt1GFP lungs stained for GFP (green), PDGFR beta (red), and PDGFR alpha (blue). Arrows indicate GFP and PDGFR beta double positive pericytes. Scale bar, 15 µm. c RT-qPCR analysis of Angpt1 expression in freshly sorted PDGFR beta + cells from P7 Yap1,Wwtr1iPCKO and control lungs. Data represents mean ± s.e.m. (n = 4 mice, two-tailed unpaired t-test). dAngpt1 expression in cultured Verteporfin (VP)-treated (48 h) and control pericytes. Data represents mean ± s.e.m. (n = 4, Welch’s t-test). e Expression of the indicated transcripts in freshly sorted CD31+ cells from P7 Yap1,Wwtr1iPCKO and control lungs. Data represents mean ± s.e.m. (n = 4 mice, NS not significant, two-tailed unpaired t-test). f–h Western blot analysis of Angpt1 protein (f; n = 2 controls and 4 mutant mice) and of total and phospho-Tie2 (pTie2) in P12 Yap1,Wwtr1iPCKO and control total lung lysates (g, n = 3 controls and 5 mutants). Molecular weight marker (kDa) is indicated. Relative quantification of signals is shown in h. Two-tailed unpaired t-test. i Scheme showing the time points of tamoxifen administration and analysis for Angpt1iPCKO mice. j, k 3D reconstruction confocal images of P12 Angpt1iPCKO and littermate control lungs stained for AQP5 (green), PDGFR beta (red), and PECAM1 (blue). Panels in k show higher magnification of PECAM1 staining. Scale bar, 50 µm (j) and 30 µm (k). l Quantitation of airspace volume in P12 Angpt1iPCKO and littermate control lung sections with 3D reconstruction surface images. Data represents mean ± s.e.m. (n = 4 mice; p < 0.0001, two-tailed unpaired t-test). m 3D reconstruction confocal images of P12 Angpt1iPCKO and littermate control lungs stained for alpha SMA (red) and tropoelastin (blue). Scale bar, 50 µm. n Quantitation of staining intensity for alpha SMA or tropoelastin shown in m. Intensity was normalized to the size of the parenchymal region. Data represents mean ± s.e.m. (n = 4 mice; NS not significant, two-tailed unpaired t-test). o Schematic summary of findings. Pulmonary pericytes regulate ECs and alveolar epithelial cells via angiocrine factors such as angiopoietin-1 and HGF Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/29934496), licensed under a CC-BY license. Not internally tested by R&D Systems.

Reactivity:

Hu, Mu

Species Glossary

Reactivity: Hu, Mu

Applications:

, , ,

Applications: , , ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

Available Formulations

Formulation	Catalog #	Availability	Price
Carrier Free	AF2720-SP

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NBP1-69753

Tie-2 Antibody - BSA Free

(3 Publications)

0.1 ml

NBP1-69753

Western Blot: Tie-2 Antibody [NBP1-69753] - Total protein from human Huvec cells and mouse stomach tissue was separated on a 7.5% gel by SDS-PAGE, transferred to PVDF membrane and blocked in 5% non-fat milk in TBST. The membrane was probed with 1.0 ug/ml anti-TIE2 in 1% non-fat milk in TBST and detected with an anti-rabbit HRP secondary antibody using chemiluminescence.

Immunohistochemistry-Paraffin: Tie-2 Antibody [NBP1-69753] - Analysis of a FFPE tissue section of mouse stomach using 1:200 dilution of Tie-2 antibody (NBP1-69753). The staining was developed using HRP labeled anti-rabbit secondary antibody and DAB reagent, and nuclei of cells were counter-stained with hematoxylin.

Western Blot: Tie-2 Antibody [NBP1-69753] - Detection of TIE2 in Huvec cell lysate

Simple Western: Tie-2 Antibody [NBP1-69753] - Simple Western lane view shows a specific band for Tie-2 in 0.5 mg/ml of HUVEC lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.

Reactivity:

Hu, Mu

Species Glossary

Reactivity: Hu, Mu

Applications:

, ,

Applications: , ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

Available Conjugates:

Conjugate	Catalog #	Availability	Size	Price
Alexa Fluor 350	NBP1-69753AF350
Alexa Fluor 405	NBP1-69753AF405
Alexa Fluor 488	NBP1-69753AF488
Alexa Fluor 532	NBP1-69753AF532
Alexa Fluor 594	NBP1-69753AF594
Alexa Fluor 647	NBP1-69753AF647
Alexa Fluor 700	NBP1-69753AF700
Alexa Fluor 750	NBP1-69753AF750
Allophycocyanin	NBP1-69753APC
Biotin	NBP1-69753B
DyLight 350	NBP1-69753UV
DyLight 405	NBP1-69753V
DyLight 488	NBP1-69753G
DyLight 550	NBP1-69753R
DyLight 594	NBP1-69753DL594
DyLight 650	NBP1-69753C
DyLight 680	NBP1-69753FR
DyLight 755	NBP1-69753IR
FITC	NBP1-69753F
HRP	NBP1-69753H
Janelia Fluor 549	NBP1-69753JF549
Janelia Fluor 646	NBP1-69753JF646
PE	NBP1-69753PE
PerCP	NBP1-69753PCP
mFluor Violet 450 SE	NBP1-69753MFV450
mFluor Violet 500 SE	NBP1-69753MFV500
mFluor Violet 610 SE	NBP1-69753MFV610

Available Formulations

Formulation	Catalog #	Availability	Price
BSA Free	NBP1-69753SS

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AF3909

Tie-2 [p Tyr1102, p Tyr1100, p Tyr1100] Antibody [Unconjugated]

(5 Publications)

100 μg

AF3909

Western blot shows lysates of NIH-3T3 mouse embryonic fibroblast cell line transfected with mouse Tie-2 untreated (-) or treated (+) with 600 ng/mL Recombinant Human Angiopoietin-1 (Catalog # 923-AN) for 5 minutes. PVDF membrane was probed with 1 µg/mL of Rabbit Anti-Human/Mouse Phospho-Tie-2 (Y1102/Y1100) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3909) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band was detected for Phospho-Tie-2 (Y1102/1100) at approximately 150 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Tie-2 phosphorylated at Y1102 was detected in immersion fixed paraffin-embedded sections of human placenta using Rabbit Anti-Human/Mouse Phospho-Tie-2 (Y1102/Y1100) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3909) at 15 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Rabbit HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS005) and counterstained with hematoxylin (blue). Lower panel shows a lack of labeling if primary antibodies are omitted and tissue is stained only with secondary antibody followed by incubation with detection reagents. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Reactivity:

Hu, Mu

Species Glossary

Reactivity: Hu, Mu

Applications:

,

Applications: ,

Host:

Rabbit Polyclonal

Host: Rabbit Polyclonal

Sizes:

Sample Size Available

Sizes: Sample Size Available

Conjugate:

Unconjugated

Available Formulations

Formulation	Catalog #	Availability	Price
Carrier Free	AF3909-SP

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