High Pressure-Accelerated Lys-C Digestion in the HUB880 Explorer: Rapid Digestion of Unreduced IgG Part II. Effect of Acetonitrile, N-Propanol, and Methanol

High Pressure-Accelerated Lys-C Digestion in the HUB880 Explorer: Rapid Digestion of Unreduced IgG Part II. Effect of Acetonitrile, N-Propanol, and Methanol

Introduction

The benefit of high pressure incubation for enhanced Lys-C digestion of unreduced IgG, and the added benefit of reagents such as urea and sodium deoxycholate, is described in separate Application Notes [4, 19]. In the current Application Note we explore the effect of several organic solvents on pressure-enhanced digestion, in order to provide the best set of starting conditions for high pressure-enhanced Lys-C digestion of disulfide-intact IgG in the presence of these reagents. These conditions are likely to be similar for digestion of other hard-to-digest proteins, such as those containing hydrophobic transmembrane domains. The current application focuses on digestion at constant high pressure (not pressure cycling) in the HUB880 Explorer.

Introduction

The benefit of high pressure incubation for enhanced Lys-C digestion of unreduced IgG, and the added benefit of reagents such as urea and sodium deoxycholate, is described in separate Application Notes [4, 19]. In the current Application Note we explore the effect of several organic solvents on pressure-enhanced digestion, in order to provide the best set of starting conditions for high pressure-enhanced Lys-C digestion of disulfide-intact IgG in the presence of these reagents. These conditions are likely to be similar for digestion of other hard-to-digest proteins, such as those containing hydrophobic transmembrane domains. The current application focuses on digestion at constant high pressure (not pressure cycling) in the HUB880 Explorer.