Accurate Glycan Analyzer
Achieves Simple, High-Sensitivity Glycan Structure Identification
Identification of Glycan Structural Isomers Using High-Precision, Multistage MSn Spectra
Described here is an example of the structural identification of pyridylaminated glycans derived from antibodies. The N-type glycans were extracted from an SDS-PAGE gel spot of human myeloma IgG1 antibody (5 ug) by PNGase F in-gel digestion. The pyridylaminated glycans were prepared by sialidase digestion, PA-labeling and purified extracted glycans.
The pyridylaminated glycans were separated using the Prominence nano liquid chromatograph, and then spotted onto a MALDI plate together with matrix solution using the Accuspot MALDI spotting device.
DHB (2,5-dihydroxybenzoic acid) was used as the matrix.
The structural identification of the pyridylaminated glycans contained in signals 2 and 4, which exhibited the same m/z value of 1,725.6, is shown above as an example. No major differences in spectral patterns could be observed in a comparison of the two MS/MS spectra. However, in MS3 spectra of the product ion at m/z 1,280.4, which was presented as first MS3 precursor ion candidate by search software via Client, relative signal intensity were different for a number of product ions at m/z 712.2 and m/z 915.3. By performing a database search with these MS/MS and MS3 spectra, glycan structure can be identified with distinguishing glycan structural isomers. The glycan structures identified from signals 1 to 8 of the mass chromatogram are shown in the table below.
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