Protein Purification-free Method of Binding Affinity Determination by Microscale ThermophoresisLyuba Khavrutskii 1,2, Joanna Yeh 1, Olga Timofeeva 3, Sergey G. Tarasov 4, Samuel Pritt 1, Karen Stefanisko 1, Nadya Tarasova 1
1Cancer and Inflammation Program, National Cancer Institute, 2Basic Science Program, SAIC-Frederick, Inc., 3Departments of Oncology and Radiation Medicine, Georgetown University Medical Center, 4Structural Biophysics Laboratory, National Cancer Institute
Microscale thermophoresis (MST) can be widely used for determination of binding affinity without purification of the target protein from cell lysates. The protocol involves overexpression of the GFP-fused protein, cell lysis in non-denaturing conditions, and detection of MST signal in the presence of varying concentrations of the ligand.
