| Assay Method Information | |
| | Fluorescence Polarization Assay |
| Description: | For competition-based fluorescence polarization assays, the ability of the test compounds to displace fluorophore-labeled peptides from their respective binding proteins was analyzed as previously described.3 Peptide sequences were: STAT1: 5-carboxyfluorescein-GY(PO3H2)DKPHVL; STAT3: 5-carboxyfluorescein-GY(PO3H2)LPQTV-NH2; STAT4: 5-carboxyfluorescein-GY(PO3H2)LPQNID-OH; STAT5a and STAT5b: 5-carboxyfluorescein-GY(PO3H2)LVLDKW; STAT6: 5-carboxyfluorescein-GY(PO3H2)VPWQDLI-OH; Lck SH2: 5-carboxyfluorescein-GY(PO3H2)EEIP. Due to protein instability, STAT2 was not analyzed. Final concentration of 5-carboxyfluorescein-labeled peptides: 10 nM. Proteins were used at the following final concentrations, which correspond to the Kd-values of the interactions with the respective fluorophore-labeled peptide: STAT1: 420 nM; STAT3: 270 nM; STAT4: 130 nM; STAT5a: 130 nM; STAT5b: 100 nM; STAT6: 310 nM; Lck SH2: 30 nM. Pipetting was carried out partly using a Biomek FX workstation (Beckman-Coulter). Pr |
| Affinity data for this assay | |
|---|---|
| If you find an error in this entry please send us an E-mail | |