Combining a Chip-Mate nESI Ion Source with Make-Up Flow Strategy for Increased System Robustness and Sensitivity in nLC-nESI-MS

摘要

Adding a post column stream of solvent to the effluent from a nano-liquid chromatography system (make-up flow strategy, MUF) allows for an optimization of the solvent composition for nano-electrospray (nESI) formation of ions and can positively influence analytical performance by effecting ionization efficiency, chromatographic peak width and nESI robustness. Here, we investigated the impact of various solvent additions post analytical column and upstream of a novel chip-based nano electrospray ion source (Chip-Mate) by using a simple splitting T and a flow rate range of 200 to 800 nL/min. A standard 6 protein digest sample was analyzed using a Waters nano Acquity nLC system, an ACE C18 75 um ID analytical column, an Advion Chip-Mate nESI ion source and a Sciex 5500 QTrap mass spectrometer using short, 30 min gradients. A selection of peptides from the mixture were analyzed for peak area and height and their dependence on the ratio of analytical flow rate vs MUF rate as well as the MUF solvent composition. Although a solvent addition effectively dilutes the analyte concentration post column and mass spectrometers are usually considered concentration dependent detectors, we found an actual increase of sensitivity with solvents containing dimethylsulfoxide (DMSO), iso-propanol (IPA) or increased vol% formic acid. So far, sensitivity gains were observed with a factor of 2-10 with some bias towards larger and hydrophobic peptides at an optimal ratio of the analytical flow to MUF of 2:1. Addition of a MUF is a simple set-up choice for any nESI system that does only require an additional nLC pumping channel with sufficient accuracy and stability in nano flow rates. MUF addition increases spray robustness, stability and longevity of a spray emitter as well as significantly increases the system sensitivity with little observed downsides so far.