Characterizing Soluble Protein Aggregates Using Native Mass Spectrometry Coupled with Temperature-Controlled Electrospray Ionization and Size-Excl usion Chromatography.

Characterization of soluble protein aggregates provides valuable information for revealing mechanisms of protein aggregation process and assessing the activity and safety of protein therapeutics. However, the noncovalent interaction, the transient nature and higher degree of structural heterogeneity of the soluble aggregation system hinders precise characterization at the molecular level. Here, we describe methods using native mass spectrometry coupled with temperature-control electrospray ionization and size-exclusion chromatography to monitor the aggregation process and profile the aggregates in detail.