Efficient Procedure for N-Glycan Analyses and Detection of Endo H-Like Activity in Human Tumor Specimens
| Autoři | |
|---|---|
| Rok publikování | 2016 |
| Druh | Článek v odborném periodiku |
| Časopis / Zdroj | JOURNAL OF PROTEOME RESEARCH |
| Fakulta / Pracoviště MU | |
| Citace | |
| www | http://pubs.acs.org/doi/abs/10.1021/acs.jproteome.6b00346 |
| Doi | http://dx.doi.org/10.1021/acs.jproteome.6b00346 |
| Obor | Analytická chemie, separace |
| Klíčová slova | Biopsy; cancer cells; EndoH; glycans; glycosylation; mass spectrometry; tumor tissue |
| Popis | Although the importance of glycosylation has been thoroughly recognized in association with a number of biological processes, efficient assessments of glycans have been hampered by both the limited size of specimens and lengthy sample preparations, particularly in clinical settings. Here we report a simple preparative method for N-glycan analyses. It involves only short one-step chloroform methanol extraction in presence or absence of water prior to PNGase F deglycosylation. The procedure was successfully applied to the investigation of N-glycans obtained from small numbers of in vitro cultured cancer cells (<= 1 x 10(5)) and to tumor tissues, including patient biopsies of small size. MALDI-MS analysis confirmed the efficient release of all N-glycan types including complex forms with poly-N-acetyllactosamine chains. In addition, nonaqueous extraction of specimens from several established cancer cell lines, as well as patient tumor tissues, yielded high-mannose glycans with one G1cNAc moiety (Man(3-9)GlcNAc), strongly suggesting preservation of enzymatic activity analogous to Endo H enzyme. In summary, the method is both a step toward the practical use of glycan profiling and a way to detect Endo H-like activity in cancer specimens. |
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