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Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis
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Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis

Article DOI: 10.3791/50541-v • 10:22 min • August 15th, 2013
August 15th, 2013 •
1,2, 1, 3, 4, 1, 1, 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

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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.

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Protein Purification-free Binding Affinity Determination Microscale Thermophoresis KD Determination Purified Protein Drug Discovery Protocol Development GFP-fused Protein Cell Lysates Non-denaturing Conditions STAT3-GFP HEK293 Cells Transcription Factor Oligonucleotides Protein Interactions Small Molecules Peptides DNA RNA
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