Detection and Quantification of Vascular Endothelial Growth Factor Receptor Tyrosine Kinases in Primary Human Endothelial Cells.
Proteins differ widely in their pattern of expression depending on organism, tissue, and regulation in response to changing conditions. In the mammalian vasculature, the endothelium responds to vascular endothelial growth factors (VEGFs) via membrane-bound receptor tyrosine kinases (VEGFRs) to modulate many aspects of vascular physiology including vasculogenesis, angiogenesis, and blood pressure. Studies on VEGFR biology are thus dependent on detecting expression levels in different cell types and evaluating how changes in protein levels correlate with changing conditions including circulating VEGF levels. Here, we present a robust immunoblot-based protocol for detecting and quantifying VEGFRs in human endothelial cells. Using internal and external standards, we can rapidly evaluate receptor copy number and assess how this is altered in response to the cellular environment.
Endothelial Cell Biology Unit, School of Molecular and Cellular Biology, University of Leeds, Leeds, LS2 9JT, UK.
Gareth W
GW
Fearnley
Stephen B
SB
Wheatcroft
Sreenivasan
S
Ponnambalam
D015153:Blotting, Western / Q000379:methods; D003720:Densitometry / Q000379:methods; D042783:Endothelial Cells / Q000378:metabolism*; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor / Q000378:metabolism*
D020794:Receptor Protein-Tyrosine Kinases; D040262:Receptors, Vascular Endothelial Growth Factor