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Anti lyve 1

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Anti-LYVE-1 is a research tool that can be used to detect and quantify the LYVE-1 protein. LYVE-1 is a receptor for the glycosaminoglycan hyaluronan and is primarily expressed on lymphatic endothelial cells. This product can be used in various research applications to study lymphatic biology and function.

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Lab products found in correlation

Anti pdgfrβ, by Cell Signaling Technology (2 mentions) Hoechst 33342, by Thermo Fisher Scientific (1 mentions) Anti mouse alexa 488, by Jackson ImmunoResearch (1 mentions) Anti goat alexa488, by Jackson ImmunoResearch (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) Anti actin, by Merck Group (1 mentions) Fibronectin, by Merck Group (1 mentions) Nocodazole, by Merck Group (1 mentions) Alexa fluor 594 coupled phalloidin, by Thermo Fisher Scientific (1 mentions) Clone 4c11, by Merck Group (1 mentions) Anti lamin b1 ab16048, by Abcam (1 mentions) Anti arp3, by Abcam (1 mentions) Latrunculin a, by Bio-Techne (1 mentions) Blebbistatin, by Bio-Techne (1 mentions) Ck666, by Bio-Techne (1 mentions) Dmi6000b inverted microscope, by Leica (1 mentions) Axio imager 2, by Zeiss (1 mentions) Cox 2, by Abcam (1 mentions) Hif 1α, by Abcam (1 mentions) Mmp 9, by Abcam (1 mentions)

Spelling variants of this product

LYVE-1 (21 citations) Goat anti-LYVE-1 (12 citations) Goat anti-mouse LYVE-1 (3 citations) Anti-LYVE-1 antibody (3 citations) Monoclonal Anti-human LYVE-1-APC (2 citations) Goat-anti-human Lyve-1 (2 citations)

10 protocols using anti lyve 1

1

Imaging and Immunoblotting of Nuclear Proteins

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For imaging the nucleus, cells were incubated with 200 ng ml−1 of Hoechst 33342 (Life Technologies) or 34580 (Invitrogen) for 30 min at 37 °C and 5% CO2.
The following antibodies were used for immunoblotting: LmnA/C (H110, Santa Cruz), anti-ArpC4 and anti-sun2 (Abcam), and anti-actin (Millipore). Antibodies were used at 1:1,000 dilution.
For immunofluorescence, we used Alexa Fluor 594-coupled phalloidin (Invitrogen; 1:400), anti-LmnAC (N18, Santa Cruz (1:50) and clone 4C11 from Sigma Aldrich (1:2,000)), anti-Lamin B1 (ab16048, Abcam (1:500)) and anti-Arp3 (Abcam; 1:200); DAPI (4,6-diamidino-2-phenylindole) for DNA staining; and secondary antibodies anti-mouse-Alexa488 and anti-Goat-Alexa488 from Jackson ImmunoResearch Laboratories were used at 1:400. Slides were mounted with custom-made Moviol.
For lymphatic vessels visualization, anti-LYVE-1 (R&D System, 1/1,000) was used.
For drug treatment CK666, latrunculin A and blebbistatin were obtained from Tocris Bioscience, Y27632 from Calbiochem, nocodazole and DMSO from Sigma-Aldrich. For silencing, Smart pool ON-TARGETplus mouse LMNA siRNA, Arpc4 siRNA and Sun1 siRNA were purchased from Thermo Fisher Scientific. Micro-channels were coated with fibronectin (Sigma) or custom-made pLL(20)-g[3.5]-PEG(2)-Rhodamin.
Thiam H.R., Vargas P., Carpi N., Crespo C.L., Raab M., Terriac E., King M.C., Jacobelli J., Alberts A.S., Stradal T., Lennon-Dumenil A.M, & Piel M. (2016). Perinuclear Arp2/3-driven actin polymerization enables nuclear deformation to facilitate cell migration through complex environments. Nature Communications, 7, 10997.
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2

Lymphatic Vessel Imaging in Mouse Ear

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Ears from C57BL/6 mice were excised and a pair of forceps was used to create a hole.The ventral and dorsal sides of the explant were separated by peeling. The ventral sheet was kept and immunostained with anti-LYVE-1 (R&D Systems) primary antibody, to mark the lymphatic vessels. After washing with media, a secondary antibody against rat (Jackson Immunoresearch, 712-166-150) was used. The ear sheet was then spread flat in a six-well plate and a PDMS block with a central hole of diameter 8 mm was placed on top of each explant. Two hundred thousand LifeAct-GFP-expressing DCs labelled with Hoechst were added in 100 μl of culture medium inside the hole. After 1 h of incubation, the ear sheet was washed with culture medium and then placed with the face on which cells were incubating against the bottom glass slide in a FluoroDish. A block of PDMS was added on top to prevent the explant from moving during imaging. Imaging was performed on an inverted confocal microscope, at 37 °C and with 5% CO2.
Thiam H.R., Vargas P., Carpi N., Crespo C.L., Raab M., Terriac E., King M.C., Jacobelli J., Alberts A.S., Stradal T., Lennon-Dumenil A.M, & Piel M. (2016). Perinuclear Arp2/3-driven actin polymerization enables nuclear deformation to facilitate cell migration through complex environments. Nature Communications, 7, 10997.
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3

Immunohistochemical Analysis of Angiogenic Markers

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The detailed immunohistochemical staining method has been described previously.12 (link) The following primary antibodies were used: rat monoclonal anti-mouse COX-1 (Cayman, Ann Arbor, MI, USA), COX-2, HIF-1α, MMP-2, and MMP-9 (Abcam, Cambridge, UK). Antibodies were diluted 1 : 50–1 : 300 and incubated overnight. Sections were observed under a light microscope (Axio Imager 2, Carl Zeiss, Oberkochen, Germany).
For immunofluorescein staining of BVs and LVs, FITC-conjugated anti-PECAM (Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-LYVE-1 (R&D Systems, Minneapolis, MN, USA), and Cy3-conjugated donkey anti-goat IgG (Jackson Immunoresearch Laboratories, West Grove, PA, USA) were used. The sections were photographed with the Leica DMI6000 B inverted microscope (Wetzlar, Germany) at × 200 magnification and automatically stitched into whole images.
Seo Y., Ji Y.W., Lee S.M., Shim J., Noh H., Yeo A., Park C., Park M.S., Chang E.J, & Lee H.K. (2014). Activation of HIF-1α (hypoxia inducible factor-1α) prevents dry eye-induced acinar cell death in the lacrimal gland. Cell Death & Disease, 5(6), e1309-.
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4

Histological Analysis of Embryonic Lung Development

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Embryos and tissues were fixed in 10% formalin overnight, dehydrated in 100% ethanol, and embedded in paraffin. 8-µm-thick sections were used for hematoxylin-eosin, periodic acid-Schiff (PAS), trichrome, and immunohistochemistry staining. The following primary antibodies were used for immunostaining: anti-LYVE1 (R&D Systems), anti-PROX1 (Abcam), anti-PECAM1 (R&D Systems; clone: 693102), anti–pro-surfactant protein C (pro-SPC; EMD Millipore), anti-Clara cell 10 (CC10; Santa Cruz Biotechnology, Inc.), anti-Podoplanin (Abcam), anti–Caveolin-1 (BD), anti-PDGFRα (Cell Signaling Technology), anti-PDGFRβ (Cell Signaling Technology), anti-SM22α (Abcam), anti-WT1 (Santa Cruz Biotechnology, Inc.), anti-Vimentin (Santa Cruz Biotechnology, Inc.), anti-Desmin (Dako), and anti-NG2 (EMD Millipore). Detailed histology procedures can be found on the University of Pennsylvania Molecular Cardiology Research Center Histology and Gene Expression Core website. Microscopic images were taken on a microscope (Eclipse 80i; Nikon) connected to a camera (DS-Ri1; Nikon). Alveolar wall thickness (averaging 50–100 measurements per embryo) and alveolar area (averaging the total alveolar area in 5–15 vision fields per embryo) were performed in Elements software (Nikon) using a 40× objective.
Jakus Z., Gleghorn J.P., Enis D.R., Sen A., Chia S., Liu X., Rawnsley D.R., Yang Y., Hess P.R., Zou Z., Yang J., Guttentag S.H., Nelson C.M, & Kahn M.L. (2014). Lymphatic function is required prenatally for lung inflation at birth. The Journal of Experimental Medicine, 211(5), 815-826.
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5

Immunohistochemical Analysis of Lymphatic Markers

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The following primary anti-human antibodies were used: sheep anti-podoplanin, goat anti- VEGFR-3, anti-LYVE-1, anti-PROX1, anti-ITGA9, anti-VEGFR-2, anti-NRP2, anti-podocalyxin and anti-GFP, all from R&D Systems, Minneapolis, MN. Mouse anti-human CD14 and anti-CD68 antibodies were from Santa Cruz Biotechnology, Dallas, TX and Thermo Fisher, Waltham, MA, respectively. Rabbit anti-acetylated histone H3 and anti-Ki-67 were from Upstate, Billerica, MA and Cell Signaling Technologies, Danvers, MA, respectively. Rabbit anti-mouse Lyve-1 and rat anti-Meca-32 antibodies were from AngioBio, Del Mar, CA, and BioXCell, West Lebanon, NH, respectively. Secondary antibodies conjugated to FITC, Cy3, DyLight 488, DyLight 549, and APC donkey anti-rabbit, anti-sheep, anti-rat, anti-mouse and anti-goat IgG were all from Jackson ImmunoResearch Laboratories (West Grove, PA).
Volk-Draper L.D., Hall K.L., Wilber A.C, & Ran S. (2017). Lymphatic endothelial progenitors originate from plastic myeloid cells activated by toll-like receptor-4. PLoS ONE, 12(6), e0179257.
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6

Meningeal CD8+ T Cell Imaging

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Anesthetized mice were euthanized, brains were sterilely dissected, and placed in ice-cold sterile PBS. Meninges within the skullcap were fixed in 4% paraformaldehyde (PFA) overnight and separated from the skullcap. Then the meninges were incubated in the block-perm buffer containing 2% normal goat serum, 1% BSA, 0.1% Triton X, and 0.05% Tween for 1 hr at room temperature with gentle rocking. The primary antibody anti-mouse CD8 (Novus, Cat# ABX-160A, 1:100), anti-lyve-1 (R&D Systems, Cat# AF2125), and the secondary antibody Alexa Fluor 488, 555 rabbit anti-mouse IgG, Alexa Fluor 555 rabbit anti-goat IgG (Life Technologies) were employed for CD8+ T cell and lymphatic vessel staining. The meninges were mounted with Prolong Gold with DAPI (Molecular Probes).
Li J., Huang D., Lei B., Huang J., Yang L., Nie M., Su S., Zhao Q, & Wang Y. (2022). VLA-4 suppression by senescence signals regulates meningeal immunity and leptomeningeal metastasis. eLife, 11, e83272.
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7

Immunohistochemistry of Lymphoid Tissues

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Immunohistochemistry was performed using a modified method of a previously published protocol (78 (link)). Briefly, freshly isolated LNs or spleens were fixed in newly prepared 4% paraformaldehyde overnight at 4°C on an agitation stage. Spleens or LNs were embedded in 4% low melting agarose (Thermo Fisher Scientific) in PBS and sectioned with a vibratome (Leica VT-1000 S) at 30 μm thickness. Thick sections were blocked in PBS containing 10% fetal calf serum, 1 mg/mL anti-Fcγ receptor (BD Biosciences), and 0.1% Triton X-100 (Sigma) for 30min at room temperature. Sections were stained overnight at 4°C on an agitation stage with the following antibodies: anti-PNAd (MECA-79, BD Biosciences) and anti-LYVE-1 (Clone 223322, R&D Systems). Stained thick sections were analyzed using a Leica SP8 confocal microscope equipped with an HC PL APO CS2 40X (NA, 1.30) oil objective (Leica Microsystem, Inc.) and images were processed with Leica LAS AF software (Leica Microsystem, Inc.) and Imaris software 9.3.1 (Bitplane AG).
Johansen K.H., Golec D.P., Huang B., Park C., Thomsen J.H., Preite S., Cannons J.L., Garçon F., Schrom E.C., Courreges C., Veres T.Z., Harrison J., Nus M., Phelan J.D., Bergmeier W., Kehrl J., Okkenhaug K, & Schwartzberg P.L. (2022). A CRISPR screen targeting PI3K effectors identifies RASA3 as a negative regulator of LFA-1–mediated adhesion in T cells. Science signaling, 15(743), eabl9169.
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8

Multicolor Immunostaining of Liver Sinusoids

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Frozen liver tissues were cut into 7 μm sections and fixed in acetone for 10 minutes and permeabilized with 0.5% Triton X-100. Slides were blocked in 10% FBS for 1 hour at room temperature before being incubated with primary antibody overnight at 4°C. Primary antibodies and stains used included anti-COL4 (Southern Biotech 1340-01; Abcam ab19808), anti-COL1 (Southern Biotech 1310-01), anti-CD34 (Abcam ab81289), anti-LYVE1 (R&D Systems AF2125), Phalloidin-TRITC (MilliporeSigma P1951), anti-COL4a1 (Chondrex 7070), anti-COL4 (Abcam ab236640), anti-PDGFRβ (Cell Signaling Technology 3169), anti-α-SMA (Abcam ab7817), and anti-Hsp47 (Novus Biologicals NBP1-97491). After incubation with fluorochrome-coupled secondary antibodies (Invitrogen, A11055, A11057, A21206, A11077, A21202, A10042) and DAPI, images were visualized on a Zeiss LSM 780 confocal microscope. 3D super-resolution images were visualized on a Zeiss LSM 980 Airyscan microscope and reconstructed and viewed with Imaris image analysis software. Colocalization analysis was done as described previously (60) . Three sinusoidal areas were selected for analysis. Fluorescence intensity peaks and ratio of green fluorescence versus red fluorescence were quantified by Zen Image Browser at 4 straight arrows as shown in the schema of hepatic sinusoids (Supplemental Figure 2D).
Gan C., Yaqoob U., Lu J., Xie M., Anwar A., Jalan-Sakrikar N., Jerez S., Sehrawat T.S., Navarro-Corcuera A., Kostallari E., Habash N.W., Cao S, & Shah V.H. (2024). Liver sinusoidal endothelial cells contribute to portal hypertension through collagen type IV-driven sinusoidal remodeling. JCI insight, 9(11).
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9

Immunostaining of Tumor Samples

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Primary tumors, axillary lymph nodes, and lungs were fixed in zinc fixative buffer (overnight, 4 °C) and 10-μm-thick paraffin sections were placed onto thermo superfrost slides (Thermo Fisher Scientific, USA). For staining, tissues were deparaffinized and blocked with 5% horse serum in 0.3% Triton X-100 for 1 h at room temperature. Then, the slides were incubated overnight (4 °C) with primary antibodies: anti-CD326 (1:100, Acris antibodies, San Diego, CA, USA; #AM33039PU-N), anti-cytokeratin (1:400, Biolegend, San Diego, CA, USA; #628602), anti-Lyve1 (1:20, R&D Systems, Minneapolis, MN, USA; #AF 2125), anti-endomucin (1:50, eBioscience, San Diego, CA, USA; #14–5851-85), anti-CD11b (1:200, Abcam, Cambridge, UK; #ab 75476), anti-F4/80 (1:100, eBioscience, San Diego, CA, USA; #14-4801). The next day, the slides were washed with 0.3% Triton X-100 in phosphate-buffered saline (PBS) and incubated with corresponding Alexa fluor secondary antibodies (room temperature, 2 h). Then, the slides were washed and mounted in Hoechst 33347 mounting medium (Sigma–Aldrich, Steinheim Germany). Images were generated using a Leica confocal microscope (Leica Microsystems, Wetzlar, Germany) and were analyzed using ImageJ software version 1.53c (NIH, MD, USA).
Kesavan R., Frömel T., Zukunft S., Brüne B., Weigert A., Wittig I., Popp R, & Fleming I. (2021). The Consequences of Soluble Epoxide Hydrolase Deletion on Tumorigenesis and Metastasis in a Mouse Model of Breast Cancer. International Journal of Molecular Sciences, 22(13), 7120.
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10

Quantifying Tumor Angiogenesis in Xenografts

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The xenograft tissue was fixed in zinc-formalin and paraffin embedded. 4 μm FFPE tissue sections were deparaffinized, dehydrated, processed with antigen retrieval buffer, and incubated with primary antibodies. For CD31 staining, citrate buffer (pH 6.0), and LYVE-1, EDTA buffer (pH 9.0) was used for antigen retrieval following the manufacturer’s protocol using a pressure cooker. The slides were then incubated with either anti-CD31 (Cell Signaling Technology, Danvers, MA, USA), or anti-LYVE1 (R&D Systems, Minneapolis, MN, USA) antibody for 24 hours at 4°C. After incubation with biotinylated secondary antibody (Vector Lab, Burlingame, CA, USA for 30 min, slides were incubated with streptavidin-biotin-peroxidase (Vector Lab, Burlingame, CA, USA). DAB substrate was used to visualize positive staining. Images were acquired with the Olympus VS200 Research Slide Scanner under 40× magnification. Microvascular density was measured using the Aperio Microvessel Analysis Algorithm (Aperio, Vista, CA, USA).
Alam M.M., Fermin J.M., Knackstedt M., Noonan M.J., Powell T., Goodreau L., Daniel E.K., Rong X., Moore-Medlin T., Khandelwal A.R, & Nathan C.A. (2023). Everolimus downregulates STAT3/HIF-1α/VEGF pathway to inhibit angiogenesis and lymphangiogenesis in TP53 mutant head and neck squamous cell carcinoma (HNSCC). Oncotarget, 14, 85-95.
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