N-glycans are important players in a variety of pathologies including different types of cancer, (auto)immune diseases, and also viral infections. Matrix-assisted laser desorption/ionization mass... Show moreN-glycans are important players in a variety of pathologies including different types of cancer, (auto)immune diseases, and also viral infections. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is an important tool for high-throughput N-glycan profiling and, upon use of tandem MS, for structure determination. By use of MALDI-MS imaging (MSI) in combination with PNGase F treatment, also spatially correlated N-glycan profiling from tissue sections becomes possible. Here we coupled laser-induced postionization, or MALDI-2, to a trapped ion mobility quadrupole time-of-flight mass spectrometer (timsTOF fleX MALDI-2, Bruker Daltonics). We demonstrate that with MALDI-2 the sensitivity for the detection of molecular [M - H](-) species of N-glycans increased by about 3 orders of magnitude. Compared to the current gold standard, the positive ion mode analysis of [M + Na](+) adducts, a sensitivity increase by about a factor of 10 is achieved. By exploiting the advantageous fragmentation behavior of [M - H](-) ions, exceedingly rich structural information on the composition of complex N-glycans was moreover obtained directly from thin tissue sections of human cerebellum and upon use of low-energy collision-induced dissociation tandem MS. In another set of experiments, in this case by use of a modified Synapt G2-S QTOF mass spectrometer (Waters), we investigated the influence of relevant input parameters, in particular pressure of the N-2 cooling gas in the ion source, delay between the two laser pulses, and that of their pulse energies. In this way, analytical conditions were identified at which molecular ion abundances were maximized and fragmentation reactions minimized. The use of negative ion mode MALDI-2-MSI could constitute a valuable tool in glycobiology research. Show less
