Fluorescence polarization assays for high-throughput screening of neuropeptide FF receptors.
We have developed the first fluorescence polarization assays of human neuropeptide FF2 receptors in 384-well microtiter plates. Assays are completed in a single well with no transfer, separation, or wash steps. The performance is suitable for high-throughput drug screening applications with regard to speed of analysis, magnitude of displaceable signal, precision, and sensitivity of various reagents. The rank order of potency of agonists and antagonists agrees well relative to the published radiometric filtration assays: DMe NPFF > NPFF > frog PP (Rana temporaria pancreatic polypeptide) > PQRFamide > BIBP 3226. The effect of highly colored compounds is very small on the polarization signal up to micromolar concentrations. The method serves as a simple and fast alternative to radioligand binding assays of antiobesity drug candidates related to NPFF receptors.
Do
Ernest U
EU
School of Agricultural Biotechnology, Seoul National University, San 56-1 Sillim-dong, Gwanak-gu, Seoul 151-742, Republic of Korea. eudo@snu.ac.kr
Choi
Gyu
G
Shin
Jaekyoon
J
Jung
Woo-suk
WS
Kim
Su-il
SI
D000818:Animals; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005454:Fluorescence Polarization / Q000379:methods*; D006801:Humans; D008024:Ligands; D010455:Peptides / Q000032:analysis*; D011896:Rana temporaria; D018013:Receptors, Neuropeptide / Q000819:agonists* / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D012680:Sensitivity and Specificity
D008024:Ligands; D010455:Peptides; D018013:Receptors, Neuropeptide; C083929:neuropeptide FF receptor