PD-1/PD-L1 binding studies using microscale thermophoresis

 

Romain Magnez, Bryan Thiroux, Solenne Taront, Zacharie Segaoula, Bruno Quesnel & Xavier Thuru

Scientific Reports
2017 vol: 7 article number: 17623 doi: 10.1038/s41598-017-17963-1

Abstract
The characterization of protein interactions has become essential in many fields of life science, especially drug discovery. Microscale thermophoresis (MST) is a powerful new method for the quantitative analysis of protein-protein interactions (PPIs) with low sample consumption. In addition, one of the major advantages of this technique is that no tedious purification step is necessary to access the protein of interest. Here, we describe a protocol using MST to determine the binding affinity of the PD-1/PD-L1 couple, which is involved in tumour escape processes, without purification of the target protein from cell lysates. The method requires the overexpression of fluorescent proteins in CHO-K1 cells and describes the optimal conditions for determining the dissociation constant. The protocol has a variety of potential applications in studying the interactions of these proteins with small molecules and demonstrates that MST is a valuable method for studying the PD-1/PD-L1 pathway.

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Topics: Biological fluorescence, Biophysical methods, Complex samples, Fusion proteins, Plant proteins, Monolith – MicroScale Thermophoresis, MST, Proteins, Publications

 

 

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Near-native, site-specific and purification-free protein labeling for quantitative protein interaction analysis by MicroScale Thermophoresis
Near-native, site-specific and purification-free protein labeling for quantitative protein interaction analysis by MicroScale Thermophoresis

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