Nt plex nanodsf grade capillaries

The stability of purified tβ₁AR was meansured by nano differential scanning fluorimetry (NanoDSF). tβ₁AR was diluted to 0.4 mg ml^–1 in protein buffer (20 mM Tris-HCl (pH 8), 0.35 M NaCl, 3 mM imidazole and 0.05% DDM). β₁AR ligands such as agonists, agonist derivatives, antagonists and partial agonists were tested at concentrations of 100 mM to measure their impact on receptor stability. Dimethylsulfoxide was added to a concentration of 5% in the final reaction volume. Control experiments involving protein alone, protein in 5% dimethylsulfoxide and ligands alone were conducted as references for measuring the stabilization effect of ligands. The protein sample and ligand were mixed at a fixed molar ratio (receptor/ligand, 1:10) and incubated for 20 min on ice before loading into NT.Plex nanoDSF-grade capillaries (NanoTemper). The melting curves of tβ₁AR were determined using Prometheus Melting Control v1.9 software (NanoTemper) by measuring the intrinsic protein fluorescence signal and its change during a temperature ramp from 20 to 95 °C at a rate of 2 °C min^–1. The melting temperature of the receptor was measured in triplicate and an average melting temperature was obtained.