Table 1.
Comparisons of labeling strategies for glycan quantitation
| Type | Method name | Pros | Cons |
|---|---|---|---|
| Metabolic incorporation/isotopic labeling | ICORA (58) | Improved reporting signal through increased O-glycan abundance, increased enrichment efficiency, optimal labeling efficiency | Only validated for O-glycans, time-restrictive, growth conditions must be carefully monitored |
| Isotopic labeling | Dimethyl labeling | Low-cost reagents, facile labeling, slight increase in glycan hydrophobicity | Limited throughput (low multiplexing capacity) |
| Isotopic labeling | Isotopic permethylation (42) | Significant improvements in glycan hydrophobicity and ionization efficiency, eight-channel multiplexing | Toxicity of iodomethane reagents |
| Isotopic labeling | Custom tags (e.g., PMP-, Gerard’s reagent P-, aniline-based etc.) | Highly customizable, effective in bespoke tagging workflows, stabilization of sialic acid residues, fixing of permanent positive charges | Concerns over labeling efficiency, need for optimization and method design |
| Mass defect | DiPyrO (66) | Greatly reduced spectral complexity, elimination of redundant sampling, precursor coisolation does not affect quantification, amine reactive tag (may be applied to glycans, peptides, and proteins) | Low multiplexing capacity (3-channels), requires higher-resolution MS1 scans, current instrumentation outperforms multiplexing capacity |
| Mass defect | mdSUGAR (67) | Labeling at glycan reducing end and on sialic acids, improved glycan fragmentation compared with commercial tags | Carbonyl-reactive tags are not as flexible in peptide and protein quantification, offer three-channel multiplexing |
| Isobaric labeling | QUANTITY (70) | Improved fragmentation and reporter ion signal, high labeling efficiency. Quaternary amin imparts permanent positive charge | Requires multistep synthesis, offers four-channel multiplexing |
| Isobaric labeling | TMT | Commercial quality control, well-characterized protocols, eight-channel multiplexing, fits within Thermo “ecosystem” | Cost-preventative |
| Isobaric labeling | SUGAR (68) | Improved b/y glycan fragment series for identification, synthesized in three high-yield steps, near 100% labeling efficiency, higher reporter ion signal for quantitation | Offers four-plex multiplexing |