Human VEGFR3/Flt-4 Antibody

VEGFR3/Flt‑4 in Human Cervical Squamous Metaplasia.

VEGFR3/Flt-4 was detected in immersion fixed paraffin-embedded sections of human cervical squamous metaplasia using Human VEGFR3/Flt-4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF349) at 15 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of Human VEGFR3/Flt-4 by Western Blot

TGF-beta 1, -beta 2 and -beta 3 reduce lymphatic marker expression in LECs.Primary human LECs were treated with 10, 20 or 30 ng/ml TGF-beta 1, -beta 2 and -beta 3 for 72 hours (a) or 100 hours (b). Untreated cells served as a control. Lysates were prepared and analysed by Western blot using antibodies specific for Lyve-1, Prox-1, VEGFR-3 or vimentin. Vinculin served as loading control. The experiment was performed twice with equivalent results. For densitometry evaluation, protein bands were analysed using the software ImageJ. Bands for the Prox-1, Lyve-1, vimentin and VEGFR-3 proteins were normalized to the corresponding loading control and are displayed as the expression level relative to the untreated control samples. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.pone.0162221), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human VEGFR3/Flt-4 by Immunocytochemistry/ Immunofluorescence

Direct interaction between PI3K p85 and VEGFR-3.A, LEC whole cell lysate was immunoprecipitated with anti-PI3K or isotype control IgG followed by Western blotting of phospho-tyrosine (p-Tyr), VEGFR-3 or PI3K. LECs were treated with vehicle control (serum-free media ‘0′), IgG control, VEGF-C (100 ng/ml, C) or VEGF-A (100 ng/ml, A) ligand for 15 minutes. Control untreated LEC whole cell lysate is indicate by ‘WCL’ and Jurkat cell lysate served as a positive control (Jurkat). B, Detection of VEGFR-3/phospho-PI3K complexes (red spots) in vitro in LECs grown in absence (B, upper left panel) or presence (B, upper middle panel) of VEGF-C (100 ng/ml) for 15 minutes; CD31 staining (green); DAPI (blue); isotype IgG control in PLA (B, upper right panel); quantification of VEGFR-3/phospho-PI3K and VEGFR-3/total-PI3K PLA signals using Olink Imaging Software (B, lower panel). Data shown as mean±s.e.m. **P<0.01 using t-test analysis. n = 3. Bar = 50 µm. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.pone.0039558), licensed under a CC-BY license. Not internally tested by R&D Systems.