Influence of C‑Trap Ion Accumulation Time on the Detectability of Analytes in IR-MALDESI MSI

Laser desorption followed by post electrospray ionization requires synchronized timing of the key events (sample desorption/ionization, mass spectrometry analysis, and sample translation) necessary to conduct mass spectrometry imaging (MSI) with adequate analyte sensitivity. In infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) MSI analyses, two laser pulses are used for analysis at each volumetric element, or voxel, of a biological sample and ion accumulation in the C-trap exceeding 100 ms is necessary to capture all sample-associated ions using an infrared laser with a 20 Hz repetition rate. When coupled to an Orbitrap-based mass spectrometer like the Q Exactive Plus, this time window for ion accumulation exceeds dynamically controlled trapping of samples with comparable ion flux by Automatic Gain Control (AGC), which cannot be used during MSI analysis. In this work, a next-generation IR-MALDESI source has been designed and constructed that incorporates a mid-infrared OPO laser capable of operating at 100 Hz and allows requisite C-trap inject time during MSI to be reduced to 30 ms. Analyte detectability of the next-generation IR-MALDESI integrated source has been evaluated as a function of laser repetition rate (100–20 Hz) with corresponding C-trap ion accumulation times (30–110 ms) in both untargeted and targeted analysis of biological samples. Reducing the C-trap ion accumulation time resulted in increased ion abundance by up to 3 orders of magnitude for analytes ranging from xenobiotics to endogenous lipids, and facilitated the reduction of voxel-to-voxel variability by more than 3-fold.