Parallel On-Target Derivatization for Mass Calibration and Rapid Profiling of N-Glycans by MALDI-TOF MS

Abstract

Glycosylation is an important post-translational modification of proteins, and abnormal glycosylation is involved in a variety of diseases. Accurate and rapid profiling of N-glycans by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) is still technically challenging and hampered mainly by mass drift of instrument, manual identification of spectrum peaks, and poor cocrystallization with traditional matrices besides low ionization efficiency of analytes. In the present study, a parallel on-target derivatization strategy (POTDS), on the basis of two rationally combined matrices, i.e., 3-hydrazinobenzoic acid plus DHB (DHB/3HBA) and quinoline-3-carbohydrazide plus DHB (DHB/Q3CH), was proposed for mass calibration and rapid detection of reducing N-glycans. Both DHB/3HBA and DHB/Q3CH show high derivatization efficiency and can improve the ionization efficiency of reducing N-glycans significantly. For mass calibration, in combination with dextrans, DHB/3HBA and DHB/Q3CH prove to be highly sensitive matrices facilitating both MS and MS2 calibration for N-glycans in dual polarities. For rapid identification, the regular mass difference observed for each N-glycan labeled with Q3CH and 3HBA respectively can eliminate the occurrence of false positives and promote automated identification of N-glycans in complex samples. For relative quantitation, the acid–base pair of DHB/Q3CH generates a concentrated cocrystallization of glycan–matrix mixtures at the edge of the droplet uniformly, exhibiting good linearity (R 2 > 0.998) and accuracy (RSD ≤ 10%). Furthermore, the established POTDS was successfully utilized to assess N-glycans of serum from HCC patients, revealing potential for biomarker discovery in clinical practice.

MALDI-2 for the Enhanced Analysis of N-Linked Glycans by Mass Spectrometry Imaging

“…Therefore, negative ion mode MS finds its application mostly in tandem MS analysis of oligosaccharides. − A hindrance for more extensively exploiting this potential is given by the rather low ion yields of MALDI for oligosaccharides in the negative ion mode. Therefore, a plethora of derivatization strategies has been developed to stabilize and neutralize the notoriously labile terminal sialic acid residues, − as well as functionalize the reducing end of the oligosaccharide with a stabilizing, often fluorescent, and permanently charged group. − Although functional, these strategies often result in complex sample preparation protocols, which are prone to the inclusion of experimentally induced measurement variations. Moreover, chemical derivatization strategies are not always transferable to tissue sections, and therefore they may not be applicable to MALDI-MSI.…”

2-Hydrazinoterephthalic Acid as a Novel Negative-Ion Matrix-Assisted Laser Desorption/Ionization Matrix for Qualitative and Quantitative Matrix-Assisted Laser Desorption/Ionization–Mass Spectrometry Analysis of N-Glycans in Peach Allergy Research

“…On the basis of our previous work, traditional organic acid matrices can be employed as the catalytic matrix, which not only improves derivatization reactivity but also enhances ionization efficiency when combined with acid-catalyzed reactive matrices. 38 In this study, four commonly used acidic matrices, including DHB, CHCA, SA, and CA, were combined with 2-HTA to create four combinational matrix systems (DHB/2-HTA, CHCA/2-HTA, SA/2-HTA, and CA/2-HTA). The derivatization was conducted with these combinational matrix systems under the same conditions (Figure 3A).…”

Advances in mass spectrometry‐based glycomics—An update covering the period 2017–2021

“…The combination of matrices can also be used to eliminate false positives. In a recent study combining 3‐hydrazinobenzoic acid plus DHB (DBH/3HBA) and quinoline‐3‐carbohydrazide plus DHB (DHB/Q3CH) to perform parallel on‐target derivatization, the two matrices with their respective tags were analyzed simultaneously [136]. DHB/3HBA and DHB/Q3CH proved to be highly sensitive matrices facilitating both MS and MS2 calibration for N ‐glycans in dual polarities, while the regular mass differences between them eliminated false positives when tested on a model carbohydrate.…”