TY - JOUR
T1 - A fluorescence polarization assay for identifying ligands that bind to vascular endothelial growth factor
AU - Peterson, Kimberly J.
AU - Sadowsky, Jack D.
AU - Scheef, Elizabeth A.
AU - Pal, Soumen
AU - Kourentzi, Katerina D.
AU - Willson, Richard C.
AU - Bresnick, Emery H.
AU - Sheibani, Nader
AU - Gellman, Samuel H.
N1 - Funding Information:
This research was supported by the National Institutes of Health (NIH) (GM56414 to S.H.G., DK50107 to E.H.B., and EY16995 to N.S.). R.C.W. was supported by the NIH and the Welch Foundation. We thank W. Seth Horne and Melissa D. Boersma for assistance with FP assay design and helpful discussions. The plate reader used for FP assays is in the University of Wisconsin–Madison W. M. Keck Center for Chemical Genomics. K.J.P. was supported in part by the NIH Chemistry–Biology Interface Training Program (T32 GM008505), J.D.S. was supported in part by a National Science Foundation (NSF) predoctoral fellowship, and S.P. was supported by a postdoctoral fellowship from the American Heart Association.
PY - 2008/7/1
Y1 - 2008/7/1
N2 - Vascular endothelial growth factor (VEGF) is a homodimeric proangiogenic protein that induces endothelial cell migration and proliferation primarily through interactions with its major receptors, VEGFR-1 and VEGFR-2. Inhibitors of one or both of these VEGF-receptor interactions could be beneficial as therapeutics for diseases caused by dysfunctional angiogenesis (e.g., cancer). Others have reported small peptides that bind to the VEGF dimer at surface regions that are recognized by the receptors. Here we report the development of a fluorescence polarization assay based on the binding to VEGF of a derivative of one of these peptides that has been labeled with BODIPY-tetramethylrhodamine (BODIPYTMR). This 384-well format assay is tolerant to dimethyl sulfoxide (DMSO, up to 4% [v/v]) and has a Z′ factor of 0.76, making it useful for identifying molecules that associate with the receptor-binding surface of the VEGF dimer.
AB - Vascular endothelial growth factor (VEGF) is a homodimeric proangiogenic protein that induces endothelial cell migration and proliferation primarily through interactions with its major receptors, VEGFR-1 and VEGFR-2. Inhibitors of one or both of these VEGF-receptor interactions could be beneficial as therapeutics for diseases caused by dysfunctional angiogenesis (e.g., cancer). Others have reported small peptides that bind to the VEGF dimer at surface regions that are recognized by the receptors. Here we report the development of a fluorescence polarization assay based on the binding to VEGF of a derivative of one of these peptides that has been labeled with BODIPY-tetramethylrhodamine (BODIPYTMR). This 384-well format assay is tolerant to dimethyl sulfoxide (DMSO, up to 4% [v/v]) and has a Z′ factor of 0.76, making it useful for identifying molecules that associate with the receptor-binding surface of the VEGF dimer.
KW - Fluorescence polarization assay
KW - High-throughput screening
KW - VEGF
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U2 - 10.1016/j.ab.2008.03.043
DO - 10.1016/j.ab.2008.03.043
M3 - Article
C2 - 18413228
AN - SCOPUS:43849105277
SN - 0003-2697
VL - 378
SP - 8
EP - 14
JO - Analytical Biochemistry
JF - Analytical Biochemistry
IS - 1
ER -