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Abstract
MALDI imaging allows for the near-cellular profiling of proteoforms directly from microbial, plant, and mammalian samples. Despite detecting hundreds of proteoforms, identification of unknowns with only intact mass information remains a distinct challenge, even with high mass resolving power and mass accuracy. To this end many supplementary methods have been used to create experimental databases for accurate mass matching, including bulk or spatially re-solved bottom-up and/or top-down proteomics. Herein we describe the application of 193 nm ultraviolet photodissociation (UVPD) for fragmentation of quadrupole isolated singly charged ubiquitin (m/z 8565) by MALDI-UVPD on an UHMR HF Orbitrap. This platform permitted the high-resolution accurate mass measurement of not just terminal fragments, but also large internal fragments. The outlined workflow demonstrates the feasibility of top-down analyses of isolated MALDI protein ions and the potential towards more comprehensive characterization of proteoforms in MALDI imaging applications.
Supplementary materials
Title
Supplementary Information
Description
Includes extended discussion about MALDI-UVPD implementation, tuning, and use with further analyses of attenuation of excimer pulse energies through optics, and several different representations of the annotated fragmentation data of various levels of microscan averaging.
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Title
Supplementary Data
Description
A zipped folder containing .RAW files corresponding to the total of 1500 microscan averages as described within this note. From an isolation acquisition with no fluence, to an experiment with 5.5 mJ pulse energy.
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