Detection of ROBO4 in HUVEC Human Cells by Flow Cytometry.
Human umbilical vein endothelial cells were stained with Mouse Anti-Human ROBO4 Monoclonal Antibody (Catalog # MAB24541, filled histogram) or isotype control antibody (Catalog # MAB0041, open histogram), followed by Allophycocyanin-conjugated Anti-Mouse IgG F(ab')2 Secondary Antibody (Catalog # F0101B).
Preparation and Storage
Reconstitute at 0.5 mg/mL in sterile PBS.
Reconstitution Buffer Available
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
ROBO4, also called magic roundabout, is a ~150 kDa glycoprotein belonging to the ROBO family (1). ROBOs are molecular guidance receptors that typically interact with Slit ligands to regulate axon guidance and neuronal migration (2). Unlike other family members, ROBO4 is mainly restricted to the vascular endothelium (1, 2). Expression in early hematopoietic progenitors is also reported (3). The human ROBO4 cDNA encodes 1012 amino acids (aa), including a 27 aa signal sequence, a 440 aa extracellular domain (ECD) containing two C2‑type Ig domains and two fibronectin type III (FNIII) domains, a transmembrane domain and an intracellular domain. ROBO4 diverges from other ROBO proteins in the number of Ig, FNIII and cytoplasmic CC domains (1, 4). Within the ECD, human ROBO4 shares 80%, 80%, 87% and 88% aa identity with mouse, rat, bovine and canine ROBO4, respectively. Vascular endothelial ROBO4 is expressed at highest levels in during development and vascular remodeling, including tumor angiogenesis (1, 2, 4‑6). It is proposed to contribute to vascular stability. Consistent with this, endogenous ROBO4 is concentrated in the vascular stalk and sprouts rather than tip cells and appears to protect newly formed blood vessels against VEGF‑induced vascular leak (6‑9). ROBO4 binding of Slit proteins has been variably reported, and when detected may be mediated by ROBO4/ROBO1 heterodimers (2, 4‑7, 9‑13). ROBO4 is also variably reported to stimulate or inhibit cell migration or filopodia formation (2, 4‑13). Effects on cell movement may be mediated through intracellular binding of WASP‑, Ras/Rac/Rho‑, Mena‑, Src‑ or Paxillin‑related proteins, all of which affect the cytoskeleton (5‑7, 10‑12). Recombinant soluble ROBO4 ECD can antagonize endothelial cell migration and in-vivo angiogenesis (13).
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