Enhanced detection of in‐gel released N‐glycans by MALDI‐TOF‐MS |
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| Authors: | Stefan Weiz Aryaline Kamalakumar Karina Biskup Véronique Blanchard |
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| Institution: | 1. Institute of Laboratory Medicine, Clinical Chemistry, and Pathobiochemistry, Charité Medical University, Berlin, Germany;2. Department of Biology, Chemistry and Pharmacy, Freie Universit?t Berlin, Berlin, Germany |
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| Abstract: | Many biologically relevant glycoproteins need to be separated on 1D‐ or 2D‐gels prior to analysis and are available in picomole amounts. Therefore, it is important to have optimized methods to unravel the glycome that combine in‐gel digestions with MALDI‐TOF‐MS. In this technical report, we investigated how the detection of in‐gel released N‐glycans could be improved by MALDI‐TOF‐MS. First, an AnchorChip target was tested and compared to ground steel target using several reference oligosaccharides. The highest signals were obtained with an AnchorChip target and D‐arabinosazone as the matrix; a LOD of 1.3 to 10 fmol was attained. Then, the effect of octyl‐β‐glucopyranoside, a nonionic detergent, was studied during in‐gel peptide‐N4‐(acetyl‐ß‐glucosaminyl) asparagine amidase F digestion of standard glycoproteins and during glycan extraction. Octyl‐β‐glucopyranoside increased the intensity and the amount of detected neutral as well as acidic N‐glycans. A LOD of under 7 pmol glycoprotein could be achieved. |
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| Keywords: | Detergent Glycan Glycoproteomics In‐gel digestion MALDI‐TOF |
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