|Detection of Human Plexin D1 by Western Blot. Western blot shows lysates of JAR human choriocarcinoma cell line, IMR‑32 human neuroblastoma cell line, and Jurkat human acute T cell leukemia cell line. PVDF membrane was probed with 0.5 µg/mL of Goat Anti-Human Plexin D1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4160) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for Plexin D1 at approximately 250 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.|
|Detection of Plexin D1 in Human peripheral blood monocytes by Flow Cytometry. Human peripheral blood monocytes were stained with Goat Anti-Human Plexin D1 Antigen Affinity‑purified Polyclonal Antibody (Catalog # AF4160, filled histogram) or control antibody (Catalog # AB‑108‑C, open histogram), followed by Phycoerythrin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0107).|
|Plexin D1 in Human Melanoma. Plexin D1 was detected in immersion fixed paraffin-embedded sections of human melanoma using Goat Anti-Human Plexin D1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4160) at 3 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-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). Specific staining was localized to plasma membranes. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.|
|Proliferation Inhibited by Semaphorin 3E and Neutralization by Human Plexin D1 Antibody. Recombinant Human Semaphorin 3E (Catalog # 3239-S3) inhibits proliferation in the HUVEC human umbilical vein endothelial cells in a dose-dependent manner (orange line). Plexin D1 mediated inhibition elicited by Recombinant Human Semaphorin 3E (5 ug/mL) is neutralized (green line) by increasing concentrations of Goat Anti-Human Plexin D1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4160). The ND50 is typically 0.5-2 ug/mL.|
Plexin D1 is a type I transmembrane glycoprotein that is the prototype of the plexin D subfamily of semaphorin receptors (1, 2). Human Plexin D1 contains a 46 amino acid (aa) signal sequence, a 1225 aa extracellular domain (ECD), a 21 aa transmembrane domain, and a 633 aa cytoplasmic domain that includes features common to other plexins (1). The human Plexin D1 ECD shares 89% identity with mouse Plexin D1, and ~84-92% aa identity based on incomplete sequences of rat, bovine, porcine and canine Plexin D1. It contains a sema domain, two plexin-semaphorin-integrin (PSI) or Met-related sequence (MRS) cysteine-rich motifs, and three glycine/proline-rich IPT/TIG domains which are immunoglobulin-like domains found in plexins, transcription factors, and the scatter factor receptors Met and Ron (1, 2). Isoforms of 1787 and 1747 aa have been sequenced; these contain a 178 aa N-terminal deletion with or without a longer alternate C-terminus (3). Like other Sema/plexin interactions, Plexin D1 interacts with Sema3C or Sema4A via neuropilins. Interaction with Sema3E, however, is direct (4). Plexin D1/Sema3E interaction mediates vascular guidance during development or angiogenesis; deletion of either molecule results in similar, profound cardiac abnormalities (4, 5). Plexin D1 is also expressed in lymphocytes, osteoblasts, the neural crest and the central nervous system during development (2, 6). In the brain, the presence of neuropilin can change Plexin D1/Sema3E interaction from an attractive to a repulsive signal (7, 8). Plexin D1 directs migration of thymocytes to the thymic medulla, probably through repulsion of Sema3E (9). Endothelial cell Plexin D1 binding to Sema4A can oppose VEGF and suppresses tumor angiogenesis, and expression of Sema3E correlates inversely with tumor metastasis, indicating that Plexin D1 is anti-metastatic in the presence of its ligands (10, 11).
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