Controlled Humidity Levels for Fine Spatial Detail Information in Enzyme-Assisted -Glycan MALDI MSI.

Investigation of the spatial distribution of -glycans in tissue specimens has emerged as a powerful tool in clinical research, in part, because altered -glycans are often a hallmark of disease progression. Mass spectrometry imaging of -glycans relies on peptide -glycanase spraying and tissue incubation for efficient release of -glycans from their carrier proteins. Unstandardized and uncontrolled incubation steps often cause significant delocalization of released -glycans, resulting in the inability to link given -glycan composition to a specific microanatomical region in the tissue. Herein, we optimized the incubation step to provide accurate and sensitive MALDI-MSI of -glycans. Specifically, we tested saturated solutions of various salts that maintain constant relative humidity in the incubation chamber. We showed that the best performance was achieved using a saturated solution of KNO that maintains an 89% RH. Under these conditions, near maximal sensitivity was achieved with the minutest ion delocalization, which we demonstrated at a 35 μm spatial resolution, where we observed six distinct spatial patterns that colocalize to distinct microanatomical compartments in a kidney nephrectomy tissue section.