L-Arginine sulfate reduces irreversible protein binding in immobilized metal affinity chromatography

•Time dependent irreversible adsorption was observed on chromatographic resins.•L-arginine can help prevent protein unfolding and reduce irreversible adsorption.•L-arginine sulfate is more effective than L-arginine chloride.•The choice of counter-ions also affects irreversible adsorption. Immobilized metal affinity chromatography (IMAC) is a powerful technique for capture and purification of relevant biopharmaceuticals in complex biological matrices. However, protein recovery can be drastically compromised due to surface induced spreading and unfolding of the analyte, leading to fouling of the stationary phase. Here, we report on the kinetics of irreversible adsorption of a protease on an IMAC resin in a timeframe ranging from minutes to several hours. This trend correlated with the thermal data measured by nano differential scanning calorimetry, and showed a time-dependent change in protein unfolding temperature. Our results highlight that ‘soft’ proteins show a strong time dependent increase in irreversible adsorption. Furthermore, commonly used cosolvents for preservation of the native protein conformation are tested for their ability to reduce fouling. Thermal data suggests that the amino acid L-arginine is beneficial to prevent unfolding, which was confirmed in batch adsorption experiments. The choice of counter-ions has to be considered when using this amino acid. These results show that L-arginine sulfate decelerates the irreversible adsorption kinetics of proteins on the IMAC stationary phase to a greater extent than L-arginine chloride.