Ultraflex 3 maldi tof ms

BOA-labeled N-glycans were directly dissolved with an equivalent volume of self-prepared ionic liquid matrix solution (100 mM α-cyano-4-hydroxycinnamic acid diethylamine salt in MeOH: H2O: DMSO: 10 mM NaOH: = 50:39:10:1), after which 2.5 μL of the sample-matrix mixture was spotted onto MTP 384 target plate (polished steel TF, Bruker Daltonics). Each sample was spotted into 4 replicates to ensure reproducibility of the experiment. After being dried to crystallize the spots, N-glycans were analyzed using Ultraflex III MALDI-TOF/MS (Bruker Daltonics, Germany) by operation in positive-ion reflector mode, typically summing 1, 000 laser shots for each spot.

Mass spectra were obtained using an Ultraflex III MALDI-TOF MS (Bruker Daltonics, Bremen, Germany) operated in positive-ion linear mode. All spectra were obtained randomly over the surface of the matrix spot. Before analysis, it was important to have the system quality controlled where 11 peptides (Bruker Daltonics, Leipzig, Germany) were used as an external standard preparation and the average molecular weight deviation was <100 μg/g; the standard preparation was used for calibration before data acquisition from samples. A coefficient of variability less than 30% indicated that the system ran well. Profile spectra were acquired from an average of 800 laser shots per sample. Peak m/z values or intensities were determined in the mass range of 1–20 kDa, focusing on mass range 1–15 kDa. Briefly, all spectra were processed by automatic baseline subtraction, peak detection, recalibration, and peak-area calculation according to the predefined settings.