Improved spatial resolution of infrared matrix‐assisted laser desorption electrospray ionization mass spectrometry imaging using a reflective objective

RATIONALE: The level of visual detail of a mass spectrometry image is dependent on the spatial resolution with which it is acquired, which is largely determined by the focal diameter in infrared laser ablation‐based techniques. While the use of mid‐IR light for mass spectrometry imaging (MSI) has advantages, it results in a relatively large focal diameter and spatial resolution. The continual advancement of infrared matrix‐assisted electrospray ionization (IR‐MALDESI) for MSI warranted novel methods to decrease laser ablation areas and thus improve spatial resolution. METHODS: In this work, a Schwarzschild‐like reflective objective was incorporated into the novel NextGen IR‐MALDESI source and characterized on both burn paper and mammalian tissue using an ice matrix. Ablation areas, mass spectra, and annotations obtained using the objective were compared against the current optical train on the NextGen system without modification. RESULTS: The effective resolution was determined to be 55 μm by decreasing the step size until oversampling was observed. Use of the objective improved the spatial resolution by a factor of three as compared against the focus lens. CONCLUSIONS: A Schwarzschild‐like reflective objective was successfully incorporated into the NextGen source and characterized on mammalian tissue using an ice matrix. The corresponding improvement in spatial resolution facilitates the future expansion of IR‐MALDESI applications to include those that require fine structural detail.