Introduce a method for coupling nano-LC with MALDI mass spectrometry, which allows to deposit the LC run on the MALDI plate in real time, and thus couple the high resolution power of nano-RP-HPLC separation directly with MALDI mass spectrometry and decouple the aquisition of MS/MS from chromatographic separation and MS cycle time.
This document details an optimized method for coupling nano-LC separation directly with MALDI mass spectrometry using the Agilent 1100 Series micro fraction collector. The system performs real-time LC run mapping on a MALDI plate, allowing for the deposition of separated fractions with online matrix addition. This approach decouples the acquisition of MS/MS analysis from the chromatographic separation cycle time.
The note outlines the experimental setup, including the use of an Agilent 1100 Series nanoflow pump, thermostatted autosampler, diode-array detector, and MSD Ion Trap XCT with AP-MALDI source. It explains the "liquid contact control mode" for reliable spotting and provides results from experiments optimizing solvent composition and plate temperature for protein identification.
| Flow rate | 800 nL/min |
|---|---|
| Flow cell volume | 80-nL |
| Matrix solution composition | 50 % 2-propanol/49 % water/1 % acetic acid |
| Optimal plate temperature | 20 °C |
The spotter ensures the capillary tip remains in contact with the surface of the deposited liquid on the MALDI target plate during the entire spotting process.
A mixture of 2 mg/mL CHCA dissolved in 50 % 2-propanol/49 % water/1 % acetic acid resulted in the best protein identification.