NMR spectroscopy allows measurements of very accurate values of equilibrium dissociation constants using chemical shift perturbation methods, provided that the concentrations of the binding partners are known with high precision and accuracy. The accuracy and precision of these experiments are improved if performed using individual capillary tubes, a method enabling full automation of the measurement. We provide here a protocol to set up and perform these experiments as well as a robust method

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... o measure peptide concentrations using tryptophan as an internal standard. Abstract 7 NMR spectroscopy allows measurements of very accurate values of equilibrium dissociation constants using 8 chemical shift perturbation methods, provided that the concentrations of the binding partners are known 9 with high precision and accuracy. The accuracy and precision of these experiments are improved if 10 performed using individual capillary tubes, a method enabling full automation of the measurement. We 11 provide here a protocol to set up and perform these experiments as well as a robust method to measure 12 peptide concentrations using tryptophan as an internal standard. 13